Construction method and application of radix comphoric oral liquid fingerprint

Abstract

The application discloses a kind of construction methods and application of lu dancheng oral liquid fingerprint, when constructing fingerprint, different batches of test solution are detected using ultra-high performance liquid chromatography, and the chromatographic column is ACQUITY Premier HSS T3, the mobile phase is 0.2wt% acetic acid aqueous solution-acetonitrile, gradient elution.The common peak separated by the fingerprint of the application is more, which provides important scientific support for evaluating the quality of lu dancheng oral liquid.The application can more objectively, comprehensively and integrally analyze and evaluate lu dancheng oral liquid by constructing fingerprint and quantitatively detecting key quality markers, provide more scientific quality control standards for industrialized production of lu dancheng oral liquid, and provide strong technical support for the consistency of quality and efficacy between different batches in the process of industrialized production of lu dancheng oral liquid.

Description

TECHNICAL FIELD

[0001] The present application relates to a kind of construction method of lu dangshen oral liquid fingerprint and the obtained fingerprint, further relates to the detection method of lu dangshen oral liquid quality marker content, further relates to lu dangshen oral liquid quality control method, belongs to the detection and analysis technical field of traditional Chinese medicine. BACKGROUND

[0002] Traditional Chinese medicine fingerprint is based on the material basis of traditional Chinese medicine, and focuses on comprehensiveness and integrity. After obtaining the comprehensive chemical information of traditional Chinese medicine by spectroscopy or chromatography, the obtained information is processed to construct the relevant fingerprint, and the quality of traditional Chinese medicine is objectively and comprehensively evaluated by similarity evaluation and chemical pattern recognition, which is of great significance to improve the overall quality control level of traditional Chinese medicine.

[0003] Lu dangshen oral liquid has the effects of tonifying qi and yin, benefiting the lung and spleen, and enhancing immunity, etc. It can help to improve the quality of life, lung function and immune function of tumor patients after radiotherapy and chemotherapy, reduce the incidence of adverse reactions, and also can reduce the degree of bone marrow suppression caused by chemotherapy in patients with lung cancer with deficiency of both qi and blood, maintain the number of white blood cells, enhance the immune function of the body and improve the quality of life of patients.

[0004] In the study of "Lu Dangshen Oral Liquid Characteristic Map and Anti-inflammatory Activity of Characteristic Peak Compound", high performance liquid chromatography technology was used to establish the characteristic map of Lu Dangshen Oral Liquid, in which 6 characteristic peaks were identified. However, the characteristic map established by this method has fewer characteristic peaks, which is not conducive to the overall quality control of Lu Dangshen Oral Liquid. In addition, there is no other report on the quality control of Lu Dangshen Oral Liquid in the prior art.

[0005] Therefore, in order to more scientifically and comprehensively evaluate the internal quality of Lu Dangshen Oral Liquid product, it is necessary to provide a detection method which can simultaneously complete the content determination and fingerprint detection of multiple components of Lu Dangshen Oral Liquid, to provide a rapid and efficient technical method for the quality control of Lu Dangshen Oral Liquid, and to reduce the workload of inspection. SUMMARY

[0006] In order to more scientifically, comprehensively and accurately evaluate the internal quality of Lu Dangshen Oral Liquid product, the present application provides a construction method of Lu Dangshen Oral Liquid fingerprint, which uses ultra performance liquid chromatography to analyze Lu Dangshen Oral Liquid. Through the screening of chromatographic conditions, especially the cooperation of chromatographic column, mobile phase and elution program, more common peaks can be separated, and the formed fingerprint can better reflect the overall quality of Lu Dangshen Oral Liquid, providing a strong guarantee for the quality control of Lu Dangshen Oral Liquid.

[0007] The Lu Codonopsis oral liquid is a Chinese patent medicine containing Lu Codonopsis pilosula, which has the effects of tonifying the middle and replenishing qi, strengthening the spleen and lungs, nourishing and strengthening the body, and enhancing the body's immune capacity. Specifically, the method for constructing the fingerprint of the Lu Codonopsis oral liquid of the present invention comprises the following steps:

[0008] (1) Using ultra-high performance liquid chromatography to test different batches of test sample solutions, and obtaining ultra-high performance liquid chromatograms of different batches of test samples;

[0009] (2) The ultra-high performance liquid chromatography (UPLC) chromatogram of the test sample was processed using the "Chinese Herbal Medicine Chromatographic Fingerprint Similarity Evaluation System" to generate a standard reference fingerprint and determine the common peaks, thus constructing the fingerprint of Ludangshen oral liquid.

[0010] Furthermore, the ultra-high performance liquid chromatography conditions include:

[0011] Column: ACQUITY Premier HSS T3 column;

[0012] Mobile phase: Mobile phase A is 0.2 wt% glacial acetic acid in water, and mobile phase B is acetonitrile;

[0013] The gradient elution program was as follows: from 0 to 11 min, the volume fraction of mobile phase A decreased from 95% to 91%; from 11 to 28 min, the volume fraction of mobile phase A decreased from 91% to 80%; from 28 to 40 min, the volume fraction of mobile phase A decreased from 80% to 71%; from 40 to 43 min, the volume fraction of mobile phase A decreased from 71% to 35%; from 43 to 48 min, the volume fraction of mobile phase A remained unchanged at 35%.

[0014] Furthermore, during ultra-performance liquid chromatography, the column temperature is 25-40°C, for example, 25°C, 30°C, 35°C, 40°C, preferably 30°C.

[0015] Furthermore, in ultra-high performance liquid chromatography, the injection volume is 1 μl-3 μl, for example, 1 μl, 2 μl, 3 μl, preferably 2 μl.

[0016] Furthermore, during ultra-performance liquid chromatography, the flow rate is 0.1-0.4 ml / min, for example, 0.1 ml / min, 0.2 ml / min, 0.3 ml / min, 0.4 ml / min, preferably 0.2 ml / min.

[0017] Furthermore, in ultra-high performance liquid chromatography, the detector used is an ultraviolet-visible spectroscopic detector, and the detection wavelength is any one of 220 nm, 250 nm, 268 nm, and 320 nm, or a combination of two or more thereof.

[0018] Further, in step (2), the chromatogram information of each test sample is introduced into the "Traditional Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System", and after multiple point correction, Mark peak matching is performed to generate a standard control fingerprint and a common mode.

[0019] Further, the step of identifying the obtained common peaks is also included. In the present application, 8 key effective components reported in the prior art in Radix Ludou- tangshen are selected as the control, namely adenosine, chlorogenic acid, syringidin, geniposide, rutin, lognside, lognol and atractylenolide III. Each control is dissolved in methanol to obtain a control solution, and then the control solutions are mixed and dissolved in methanol to obtain a mixed control solution. The mixed control solution is detected under the above high performance liquid chromatography conditions to obtain a mixed control chromatogram. The common peaks in the constructed Radix Ludou-tangshen oral liquid fingerprint are identified according to the mixed control chromatogram, and 8 chromatographic peaks are identified, peak 1 is adenosine, peak 7 is chlorogenic acid, peak 8 is syringidin, peak 9 is geniposide, peak 14 is rutin, peak 18 is lognside, peak 21 is lognol, and peak 25 is atractylenolide III.

[0020] Further, different batches of Radix Ludou-tangshen oral liquid are selected as test samples to ensure the accuracy of the fingerprint. The preparation method of the test sample solution is as follows: 2 ml of Radix Ludou-tangshen oral liquid of different batches is taken and placed in a 5 ml volumetric flask. After adding a solvent, ultrasonic is performed for 5-30 min, and then the volume is adjusted to the mark, shaken uniformly, and centrifuged at 12000 r / min for 10 min. The supernatant is filtered to obtain the test sample solution. The solvent is methanol, 50wt% methanol aqueous solution or water, and methanol is preferred.

[0021] The present application also provides a Radix Ludou-tangshen oral liquid fingerprint constructed according to the above method. The fingerprint determines 18 common peaks at a detection wavelength of 250 nm, 16 common peaks at a detection wavelength of 268 nm, 8 common peaks at a detection wavelength of 220 nm, and 6 common peaks at a detection wavelength of 320 nm.

[0022] The present application also provides a method for detecting the content of quality markers of Radix Ludou-tangshen oral liquid. The quality markers are chlorogenic acid, syringidin, geniposide, lognside, lognol and atractylenolide III. The test sample of Radix Ludou-tangshen oral liquid is detected by ultra-high performance liquid chromatography, and then the content of each quality marker is obtained by external standard method. The ultra-high performance liquid chromatography conditions are the same as described above.

[0023] Further, the method for detecting the content of each quality marker includes the following steps:

[0024] (1) Each quality marker is prepared into a solution with methanol, and then the quality marker solutions are mixed and fixed to volume with methanol to obtain a mixed quality marker solution. The mixed quality marker solution is diluted with methanol to a series of different concentrations to obtain a series of reference solution;

[0025] (2) A series of reference solutions were tested by ultra-high performance liquid chromatography (UPLC) under the same UPLC conditions as described above. The peak area of ​​each mass marker was obtained, and a standard curve of the concentration and peak area of ​​each mass marker was drawn to obtain the regression equation of each mass marker.

[0026] (3) The test solution of Ludangshen oral liquid was tested by ultra-high performance liquid chromatography. The ultra-high performance liquid chromatography conditions were the same as those described above. The peak area of ​​each mass marker was obtained and substituted into the regression equation to calculate the content of each mass marker.

[0027] Furthermore, when detecting chlorogenic acid, the best detection wavelength is 320nm; when detecting syringin, codonopsis pilosula, and codonopsis pilosula alcohol, the best detection wavelength is 268nm; when detecting geniposide, the best detection wavelength is 250nm; and when detecting atractylodes lactone III, the best detection wavelength is 220nm.

[0028] Furthermore, the preparation method of the test solution of Lu Codonopsis Oral Liquid is as follows: 2 ml of Lu Codonopsis Oral Liquid is placed in a 5 ml volumetric flask, solvent is added, ultrasonicated for 5-30 minutes, the volume is adjusted to the scale, shaken, centrifuged at 12,000 rpm for 10 minutes, and the supernatant is filtered to obtain the test solution. The solvent is methanol, 50 wt% methanol aqueous solution or water, preferably methanol.

[0029] The present invention also provides a method for constructing the fingerprint of the Ludangshen oral liquid, or / and a method for detecting the content of the quality marker of the Ludangshen oral liquid, or / and the application of the fingerprint of the Ludangshen oral liquid in quality detection, quality evaluation, and / or quality control of the Ludangshen oral liquid. By using the fingerprint and the quantitative detection of the quality markers, either alone or in combination, the authenticity and quality of the Ludangshen oral liquid can be effectively determined, and its quality can be evaluated. The content of the active ingredients of different batches and different Ludangshen oral liquids from the same batch can also be effectively controlled, thereby ensuring the stability of the product quality and efficacy. The present invention provides a rapid and efficient technical method for quality control of the Ludangshen oral liquid, reducing the inspection workload.

[0030] The present invention has the following beneficial effects:

[0031] 1. The present application improves the accuracy of separation by screening chromatographic conditions, especially the synergistic collocation of chromatographic column, mobile phase and gradient elution program, the obtained fingerprint has more common peaks, solves the problem of less common peaks and difficult separation of the prior art, and provides important scientific support for evaluating the quality of Lu Dangshen oral liquid.

[0032] 2. The present application can more objectively, comprehensively and integrally analyze and evaluate Lu Dangshen oral liquid through fingerprint construction and quantitative detection of key effective components, provides more scientific quality control standards for industrialized production of Lu Dangshen oral liquid, and provides strong technical support for the consistency of quality and efficacy of Lu Dangshen oral liquid in different batches in the industrialized production process. BRIEF DESCRIPTION OF DRAWINGS

[0033] Figure 1a Fingerprint of 15 batches of Lu Dangshen oral liquid samples in Example 1 and the generated standard control fingerprint (250nm).

[0034] Figure 1b Fingerprint of 15 batches of Lu Dangshen oral liquid samples in Example 1 and the generated standard control fingerprint (268nm).

[0035] Figure 1c Fingerprint of 15 batches of Lu Dangshen oral liquid samples in Example 1 and the generated standard control fingerprint (220nm).

[0036] Figure 1d Fingerprint of 15 batches of Lu Dangshen oral liquid samples in Example 1 and the generated standard control fingerprint (320nm).

[0037] Figure 2a Fingerprint of the mixed control at 250nm.

[0038] Figure 2b Fingerprint of the mixed control at 268nm.

[0039] Figure 2c Fingerprint of the mixed control at 220nm.

[0040] Figure 2d Fingerprint of the mixed control at 320nm.

[0041] Figure 3 Standard fingerprint of Lu Dangshen oral liquid (R) (18 common peaks at 250nm).

[0042] Figure 4 Fingerprint of 8 common peaks of Lu Dangshen oral liquid standard fingerprint (250nm).

[0043] Figure 5This is the precision test result of Example 1 (250nm).

[0044] Figure 6 The stability test results of Example 1 (250 nm) are shown.

[0045] Figure 7 This is the repeatability test result of Example 1 (250 nm).

[0046] Figure 8 This is the elongation test result of Example 1 (250 nm).

[0047] Figure 9 This is the standard curve diagram of the six quality markers of Ludangshen Oral Liquid.

[0048] Figure 10 UPLC chart of Ludangshen oral liquid test sample at a wavelength of 210-360nm.

[0049] Figure 11 Ultra-high performance liquid chromatograms (250 nm) of different chromatographic columns.

[0050] Figure 12 Ultra-high performance liquid chromatograms (250 nm) with different mobile phase compositions.

[0051] Figure 13 Ultra-high performance liquid chromatograms (250 nm) with different elution programs.

[0052] Figure 14 Ultra-high performance liquid chromatograms (250 nm) at different column temperatures.

[0053] Figure 15 Ultra-high performance liquid chromatograms (250 nm) with different injection volumes.

[0054] Figure 16 Ultra-high performance liquid chromatograms (250 nm) at different flow rates.

[0055] Figure 17 Ultra-high performance liquid chromatograms (250 nm) of different extraction methods.

[0056] Figure 18 Ultra-high performance liquid chromatograms (250 nm) of different extraction solvents.

[0057] Figure 19 Ultra-high performance liquid chromatograms (250 nm) of different sampling amounts. DETAILED DESCRIPTION

[0058] The following is a clear and complete description of the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0059] The instruments used are as follows:

[0060]

[0061] The controls are as follows:

[0062]

[0063] The test products used in Examples 1 and 2 are as follows:

[0064]

[0065] The reagents used are as follows:

[0066]

[0067] Example 1 UPLC fingerprint methodological verification of Ludangshen oral liquid

[0068] 1. Chromatographic conditions:

[0069] Chromatographic column: ACQUITY Premier HSS T3 (2.1 × 100 mm, 1.8 μm); detection wavelengths: 250 nm, 268 nm, 220 nm, 320 nm; column temperature: 30°C; flow rate: 0.2 ml / min; injection volume: 2 μl; mobile phase A: 0.2 wt% glacial acetic acid in water; mobile phase B: acetonitrile.

[0070] Gradient elution program:

[0071]

[0072] 2. Construction of fingerprint

[0073] (1) Solution preparation:

[0074] Preparation of reference substance stock solution: Accurately weigh appropriate amounts of adenosine, chlorogenic acid, syringoside, geniposide, rutin, codonopsis arginine, codonopsis arginine alcohol, and atractylodes lactone III reference substances, and add methanol to each of them to prepare a reference substance stock solution containing 0.159 mg of adenosine, 0.168 mg of chlorogenic acid, 1.178 mg of syringoside, 1.418 mg of geniposide, 0.830 mg of rutin, 1.075 mg of codonopsis arginine, 0.975 mg of codonopsis arginine alcohol, and 0.331 mg of atractylodes lactone III per 1 ml.

[0075] Preparation of mixed reference solution: Accurately pipette the reference stock solution into a volumetric flask, add methanol to make up to volume, and obtain a mixed reference solution containing adenosine, chlorogenic acid, syringin, geniposide, rutin, codonopsis pilosula, codonopsis arginol, and atractylodes lactone III, wherein the concentration of adenosine is 15.90 μg / ml, the concentration of chlorogenic acid is 2.69 μg / ml, the concentration of syringin is 37.64 μg / ml, the concentration of geniposide is 22.69 μg / ml, the concentration of rutin is 13.26 μg / ml, the concentration of codonopsis pilosula, the concentration of codonopsis arginol is 17.47 μg / ml, the concentration of codonopsis arginol is 12.68 μg / ml, and the concentration of atractylodes lactone III is 5.30 μg / ml.

[0076] Preparation of test solution for each batch: accurately measure 2 ml of each batch of Ludangshen oral liquid, place it in a 5 ml volumetric flask, add methanol and ultrasonicate for 5 minutes, dilute to the scale, shake well, centrifuge at 12000 rpm for 10 minutes, filter the supernatant, and take the filtrate as the test solution.

[0077] (2) Ultra-high performance liquid chromatography detection and data processing:

[0078] The mixed reference solution and each test solution were tested in turn according to the chromatographic conditions of this embodiment to obtain the mixed reference chromatogram and the test sample chromatogram. The mixed reference chromatogram and the test sample chromatogram in the "CDF" format were imported into the 2012 edition of the "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System" recommended by the Pharmacopoeia Committee for data processing. The chromatographic peak of the test sample 1 (sample number S1) was used as the reference spectrum. The average method was used, and the time window width was set to 0.1. After multi-point correction, Mark peak matching was performed to generate a standard control fingerprint spectrum (R) and a common pattern. The control fingerprint spectrum (R) is the constructed Ludangshen oral liquid fingerprint spectrum. The standard control fingerprint spectrum (R) and the fingerprint spectrum of each test sample are shown in Figures 1a to 1d , the fingerprint of mixed reference substance is shown in Figures 2a to 2d . Among them, there are 18 common peaks in the standard control fingerprint (R) at 250nm, namely peak 1, peak 3, peak 5, peak 8, peak 9, peak 10, peak 11, peak 12, peak 14, peak 16, peak 17, peak 18, peak 19, peak 20, peak 21, peak 22, peak 23, and peak 25; there are 16 common peaks at the detection wavelength of 268nm, namely peak 1, peak 2, peak 3, peak 4, peak 6, peak 8, peak 9, peak 11, peak 13, peak 14, peak 17, peak 18, peak 19, peak 20, peak 21, and peak 24; there are 8 common peaks at the detection wavelength of 220nm, namely peak 6, peak 8, peak 9, peak 11, peak 13, peak 14, peak 18, and peak 25; there are 6 common peaks at the detection wavelength of 320nm, namely peak 3, peak 7, peak 8, peak 13, peak 14, and peak 15.

[0079] Enlarged view of the standard control fingerprint chromatogram R (250 nm) of Lu Dangshen oral liquid Figure 3 , with peak 18 as the reference peak, the relative retention times of each common peak were as follows: peak 1: 0.0917, peak 3: 0.1679, peak 5: 0.2138, peak 8: 0.4332, peak 9: 0.5445, peak 10: 0.6032, peak 11: 0.6160, peak 12: 0.7216, peak 14: 0.8258, peak 16: 0.8815, peak 17: 0.9119, peak 18: 1.0000, peak 19: 1.0404, peak 20: 1.1320, peak 21: 1.1483, peak 22: 1.1754, peak 23: 1.2004, peak 25: 1.3625. According to the fingerprint chromatogram of the mixed control sample, the common peaks of Lu Dangshen oral liquid were identified, and 8 chromatographic peaks were identified, peak 1 was adenosine, peak 7 was chlorogenic acid, peak 8 was syringin, peak 9 was geniposide, peak 14 was rutin, peak 18 was notoginsenoside, peak 21 was notoginsenoside, and peak 25 was atractylenolide III, as shown in Figure 4 (250 nm).

[0080] The chromatograms of 15 batches of Lu Dangshen oral liquid samples were compared with the similarity of the standard control fingerprint chromatogram R at different detection wavelengths, and the results are shown in Table 1:

[0081] Table 1: Similarity results of 15 batches of Lu Dangshen oral liquid chromatogram and standard control fingerprint chromatogram (R)

[0082]

[0083] From the similarity calculation results, it can be seen that the similarity of the fingerprint chromatograms of different batches of Lu Dangshen oral liquid and the standard control fingerprint chromatogram (R) of Lu Dangshen oral liquid at each wavelength is higher than 0.9, which indicates that the similarity is good, and the UPLC control fingerprint chromatogram "R" obtained by fitting can comprehensively represent the information of the effective components in Lu Dangshen oral liquid.

[0084] 3. Verification content

[0085] 3.1 Precision

[0086] (1) Experimental steps

[0087] Preparation of test solution: accurately take 2 ml of Lu Dangshen oral liquid (batch number 20240123) and place it in a 5 ml volumetric flask. Add methanol and ultrasonic for 5 min, then dilute to the mark, shake well, centrifuge at 12000 rpm for 10 min, filter the supernatant, and take the filtrate as the test solution.

[0088] Ultra-high performance liquid chromatography was performed six times in succession according to the chromatographic conditions of step 1 of this embodiment. The relative retention time and relative peak area of ​​each common peak at four different detection wavelengths of 250 nm, 268 nm, 220 nm and 320 nm were recorded and analyzed. Taking the detection wavelength of 250 nm as an example, the detection results are shown in Tables 2, 3 and Attached. Figure 5 shown.

[0089] (2) Experimental results

[0090] Table 2: Precision experiment - relative retention time (250nm, with peak 18 as the reference peak)

[0091]

[0092] Table 3: Precision experiment - relative peak area (250nm, with peak 18 as the reference peak)

[0093]

[0094] (3) Conclusion

[0095] The results showed that the relative retention times of the chromatographic peaks of the test solution of Ludangshen oral solution at each detection wavelength were basically consistent (RSD≤5%), the relative peak areas of the peaks were basically consistent (RSD≤5%), and the precision test met the requirements.

[0096] 3.2 Stability

[0097] (1) Experimental steps

[0098] The test solution was prepared according to the method under "3.1" of Example 1 above and stored at room temperature. Ultra-high performance liquid chromatography was performed at 0 hours, 3 hours, 6 hours, 9 hours, 12 hours, and 24 hours according to the chromatographic conditions of step 1 of this example. The relative retention time and relative peak area of ​​each common peak at four different detection wavelengths of 250 nm, 268 nm, 220 nm, and 320 nm were recorded and analyzed. Taking the detection wavelength of 250 nm as an example, the test results are shown in Tables 4, 5, and Attached. Figure 6 shown.

[0099] (2) Experimental results

[0100] Table 4: Stability test - relative retention time (250nm, with peak 18 as the reference peak)

[0101]

[0102] Table 5: Stability test - relative peak area (250nm, with peak 18 as the reference peak)

[0103]

[0104] The results showed that the relative retention times of the chromatographic peaks of the test solution of Lu Dangshen oral liquid at each detection wavelength were basically consistent (RSD≤5%), and the relative peak areas of the peaks were basically consistent (RSD≤5%), indicating that the test solution was stable within 24 hours.

[0105] 3.3 Repeatability

[0106] (1) Experimental steps

[0107] Prepare the test solution according to the method under "3.1" of step 1 above, and operate in parallel for 6 times. Perform ultra-high performance liquid chromatography detection 6 times in succession according to the chromatographic conditions of step 1 of this example. Record and analyze the relative retention time and relative peak area of ​​each common peak at four different detection wavelengths of 250nm, 268nm, 220nm and 320nm. Taking the detection wavelength of 250nm as an example, the detection results are shown in Tables 6, 7 and Appendix Figure 7 shown.

[0108] (2) Experimental results

[0109] Table 6: Repeatability experiment - relative retention time (250nm, with Peak 18 as the reference peak)

[0110]

[0111] Table 7: Repeatability experiment - relative peak area (250nm, with peak 18 as the reference peak)

[0112]

[0113] The results showed that the relative retention times of the chromatographic peaks of the test solution of Ludangshen oral solution at each detection wavelength were basically consistent (RSD≤5%), and the relative peak areas of the peaks were basically consistent (RSD≤5%), and the repeatability experiment met the requirements.

[0114] 3.4 Extended experiment

[0115] (1) Experimental steps

[0116] The test solution was prepared according to the method under "3.1" of Example 1 above, and ultra-high performance liquid chromatography (UPLC) was performed for 90 minutes under the chromatographic conditions of step 1 of this example, and the chromatogram was recorded. Pure methanol was used as a control.

[0117] (2) Experimental results

[0118] The test results are as attached Figure 8 As shown in the attached figure, the peak that appears 48 minutes after injection is the solvent peak, and the original chromatographic conditions can cover relatively complete chromatographic information.

[0119] Example 2 Quantitative Detection of Quality Markers in Ludangshen Oral Liquid

[0120] 1. Chromatographic conditions:

[0121] The chromatographic conditions were the same as those in Example 1, except that chlorogenic acid was detected at a wavelength of 320 nm, syringin, codonopsis pilosula, and codonopsis pilosula alcohol were detected at a wavelength of 268 nm, geniposide was detected at a wavelength of 250 nm, and atractylodes lactone III was detected at a wavelength of 220 nm.

[0122] 2. Methodological Investigation

[0123] 2.1 Determination of standard curve and regression equation

[0124] (1) Solution preparation

[0125] Preparation of linear stock solutions: Accurately weigh chlorogenic acid, syringin, geniposide, codonopsis arginine, codonopsis arginine alcohol, and atractylodes lactone III reference standards. Add methanol to each to create linear stock solutions containing 0.168 mg of chlorogenic acid, 1.178 mg of syringin, 1.418 mg of geniposide, 1.075 mg of codonopsis arginine, 0.975 mg of codonopsis arginine alcohol, and 0.331 mg of atractylodes lactone III per 1 ml. Preparation of mixed mass marker solution: Accurately measure appropriate amounts of chlorogenic acid linear stock solution, syringin linear stock solution, geniposide linear stock solution, codonopsis arginine linear stock solution, codonopsis arginine alcohol linear stock solution, and atractylodes lactone III linear stock solution into a 10 ml volumetric flask, dilute to the mark with methanol, and shake well to obtain a mixed mass marker solution. Preparation of mixed reference solution: Dilute the mixed reference solution with methanol to a series of different concentrations to obtain a mixed reference solution.

[0126] (2) Chromatographic detection

[0127] According to the chromatographic conditions of step 1 of this example, a series of mixed reference solutions with different concentrations were subjected to ultra-high performance liquid chromatography to obtain the peak areas of different mass markers. Standard curves of different mass markers were drawn with mass concentration as the abscissa (X) and peak area as the ordinate (Y). Then, the linear regression equation and correlation coefficient (R) of each mass marker were calculated. 2 ), the results are shown in Table 8 and Figure 9 .

[0128] Table 8: Linear relationship results of 6 quality markers

[0129]

[0130] As shown in Table 8, all standard curves showed good linear relationship within the linear range (R 2 All ≥0.9993). This shows that the detection of ultra-high performance liquid chromatography has linear stability.

[0131] 2.2 Sample recovery

[0132] Accurately measure the reference standard linear stock solution from "2.1" in Example 2 above into a 5 ml volumetric flask and dilute to the mark with methanol. This results in a reference standard solution containing 20.16 μg chlorogenic acid, 212.04 μg syringin, 68.064 μg geniposide, 139.75 μg codonopsis arginine, 29.25 μg codonopsis arginine alcohol, and 33.1 μg atractylodes lactone III per 1 ml. Accurately measure 1 ml of Ludangshen Oral Liquid (Batch No. 20240123) into a 5 ml volumetric flask. Add the reference standard solution to the sample, ensuring a mass ratio of approximately 1:1 between each quality marker in the reference standard and each quality marker in the sample. After sonication with methanol for 5 minutes, bring the solution to volume, shake well, and centrifuge at 12,000 rpm for 10 minutes. Filter the supernatant and use it as the 100% recovery solution (prepare 6 portions using the same method). Inject the sample according to the chromatographic conditions of step 1 of this example, and calculate the content. The results showed that the recovery rates of chlorogenic acid, syringin, geniposide, codonopsis pilosula, codonopsis pilosula alcohol, and atractylodes lactone III in the samples were all between 95% and 105%, and the RSDs of the six recoveries were all less than 2%, meeting the requirements.

[0133] 2.3 Precision

[0134] Accurately measure the reference substance linear stock solution described in "2.1" of Example 2 above into a 2 ml volumetric flask. Dilute to volume with methanol to prepare a mixed reference substance solution containing 2.69 μg of chlorogenic acid, 37.64 μg of syringin, 22.69 μg of geniposide, 17.47 μg of codonopsis pilosula, 12.68 μg of codonopsis pilosulphinol, and 5.30 μg of atractylodes lactone III per 1 ml. The mixed reference substance solution was subjected to ultra-high performance liquid chromatography (UPLC) six times according to the chromatographic conditions of step 1 of this example. The chromatograms were recorded, and the peak areas of the six mass markers were calculated. The results showed that the RSDs of the peak areas for chlorogenic acid, syringin, geniposide, codonopsis pilosula, codonopsis pilosulphinol, and atractylodes lactone III were all less than 2%, demonstrating the high precision of the UHPLC assay.

[0135] 2.4 Stability

[0136] The test solution was prepared according to the method under "3.2" of Example 1 above, and ultra-performance liquid chromatography analysis was performed according to the chromatographic conditions of step 1 of this example at 0 hours, 3 hours, 6 hours, 9 hours, 12 hours, and 24 hours. The results showed that the RSD of the peak areas of the six quality markers was less than 2% at different storage times. This shows that the stability of the Ludangshen oral liquid test solution within 24 hours is good.

[0137] 2.5 Repeatability

[0138] Six test sample solutions were prepared according to the method under "3.3" of Example 1 above and subjected to ultra-performance liquid chromatography analysis according to the chromatographic conditions of step 1 of this example. The contents of the six mass markers were calculated. The results showed that the RSDs of the contents of the six mass markers in the six test samples were all less than 2%, indicating good reproducibility.

[0139] 2.6 Determination of Quality Markers in 15 Batches of Ludangshen Oral Liquid

[0140] (1) Experimental steps

[0141] Preparation of reference solution: Accurately measure the reference standard linear stock solution described in "2.1" of Example 2 above into a 5 ml volumetric flask and dilute to the mark with methanol to prepare a mixed reference solution containing 2.69 μg chlorogenic acid, 37.70 μg syringin, 22.69 μg geniposide, 17.20 μg codonopsis arginine, 12.48 μg codonopsis arginine alcohol, and 5.30 μg atractylodes lactone III per 1 ml. Prepare the test solution according to the method described in "3.1" of Example 1 above. Perform ultra-performance liquid chromatography (UPLC) analysis according to the chromatographic conditions of step 1 of this example and calculate the contents of chlorogenic acid, syringin, geniposide, codonopsis arginine, codonopsis arginine alcohol, and atractylodes lactone III in Lu Dangshen oral solution.

[0142] (2) Experimental results

[0143] Table 9: Results of determination of 6 quality markers

[0144]

[0145] As can be seen from the table, each ml of Lu Codonopsis oral liquid contains chlorogenic acid, syringin, geniposide, codonopsis arginine, codonopsis arginine alcohol and atractylodes lactone III: 7.2409-13.6693μg, 54.2260-129.2290μg, 18.5809-38.3705μg, 73.5731-207.4336μg, 7.4856-29.9791μg and 12.9757-37.4842μg respectively.

[0146] Example 3 Screening of chromatographic conditions

[0147] 1. Preparation of test solution: Prepare the test solution according to the method under "3.1" of Example 1.

[0148] 2. Chromatographic conditions: the same as those in step 1 of Example 1.

[0149] 3.1. Determination of detection wavelength: According to the chromatographic conditions of step 2 of this example, the above test sample was added to the ultra-high performance liquid chromatograph, and the full wavelength range of 210-360 nm was selected for detection based on the literature and the chromatographic peak response. The results are as follows Figure 10 As shown, the results show that the chromatograms obtained at wavelengths of 220nm, 250nm, 268nm and 320nm have good overall chromatographic peak separation, a large number of peaks, especially the chromatographic peak separation of key quality markers is high and the peak shape is symmetrical. Therefore, 220nm, 250nm, 268nm and 320nm were determined as the detection wavelengths.

[0150] 3.2. Screening of chromatographic columns: The chromatographic columns in the chromatographic conditions of step 2 of this example were replaced with three different chromatographic columns, and then the test sample was detected according to the chromatographic conditions of step 2 of this example, with a detection wavelength of 250 nm. The chromatographic columns were: ACQUITY Premier HSS T3 chromatographic column (2.1×100 mm, 1.8 μm), ACQUITY BEH chromatographic column (2.1×100 mm, 1.7 μm), and Acclaim™ RSLC 120 C18 chromatographic column (2.1×100 mm, 2.2 μm). The test results are as follows: Figure 11 As shown. Comparing the chromatograms of different columns under the same elution conditions, when ACQUITY BEH was used for elution, syringin at 11 minutes could not be well separated, and the common peak at 36 minutes had poor resolution. When Acclaim™ RSLC 120C18 was used for elution, the resolution of the geniposide peak at 17 minutes was poor, the common peaks at 19-22 minutes could hardly be well separated, and no common peaks were obtained at 35-42 minutes. When ACQUITY Premier HSS T3 was used, the fingerprint of the test solution showed good resolution of each peak, and the peak response was better than that of other columns. Therefore, ACQUITY Premier HSS T3 was selected as the column for the fingerprint determination of this oral solution.

[0151] 3.3. Mobile Phase Screening: The mobile phases in the chromatographic conditions of step 2 of this example were replaced with six different mobile phases. The test samples were then detected according to the chromatographic conditions of step 2 of this example at a detection wavelength of 250 nm.

[0152] The mobile phase is as follows:

[0153]

[0154] The experimental results are as follows Figure 12As shown. Comparing the chromatograms of different mobile phases under the same elution conditions, when acetonitrile-water is used as the mobile phase for elution, the chlorogenic acid peak separation at 12-14 minutes is poor and cannot be well separated. Compared with the elution results of other mobile phase compositions, more separated peaks cannot be obtained at 19-22 minutes and 35-43 minutes. When acetonitrile-0.1wt% glacial acetic acid aqueous solution is used as the mobile phase for elution, chlorogenic acid cannot be well separated at 13-14 minutes, and the common peak at 21 minutes cannot be well separated. When acetonitrile-0.1wt% formic acid aqueous solution is used as the mobile phase for elution, chlorogenic acid cannot be well separated at 13-14 minutes, and the common peak at 37 minutes cannot be well separated. When acetonitrile-0.1wt% phosphoric acid aqueous solution is used as the mobile phase for elution, chlorogenic acid cannot be well separated at 13-14 minutes. When acetonitrile-0.3wt% glacial acetic acid aqueous solution is used as the mobile phase for elution, syringin cannot be well separated at 8 minutes. When acetonitrile-0.2wt% glacial acetic acid aqueous solution was used, the fingerprint of the test solution showed that the peaks were well separated, and the response of the qualitative peak was better than that of other mobile phase compositions. Therefore, acetonitrile-0.2wt% glacial acetic acid aqueous solution was selected as the mobile phase composition used for the fingerprint determination of this oral liquid.

[0155] 3.4. Screening of elution program: Replace the elution program in the chromatographic conditions of step 2 of this embodiment, and then detect the test sample according to the chromatographic conditions of step 2 of this embodiment to screen the elution program. The detection wavelength is 250 nm.

[0156] The elution program is as follows:

[0157] Elution procedure 1. Mix mobile phase A and mobile phase B in a volume ratio of 75:25, and then perform isocratic elution with an elution time of 40 minutes.

[0158] Elution procedure 2:

[0159]

[0160] Elution procedure 3:

[0161]

[0162] Elution procedure 4:

[0163]

[0164] Elution procedure 5:

[0165]

[0166] The experimental results are as follows Figure 13As shown. As can be seen from the figure, when elution program 1 is used for isocratic elution, no obvious characteristic peaks appear. When elution program 2 is used for elution, the 10-12min chromatographic peaks aggregate, and it is basically impossible to achieve good separation. When elution program 3 is used, there is a common peak accumulation before 30min, and it is basically impossible to achieve good separation. When elution program 4 is used, the chromatographic peaks aggregate at 35-45min, and the total elution time is 60min, which is relatively long. When elution program 5 is used, the fingerprint of the test solution of the test sample shows that the separation of each peak is good, and the response of the peak is better than that of other elution conditions. Therefore, elution program 5 is selected as the elution condition used for the determination of the fingerprint of this oral liquid.

[0167] 3.5. Screening of column temperature: Chromatographic conditions: mobile phase B is acetonitrile, mobile phase A is 0.1wt% glacial acetic acid aqueous solution, column temperature is 25℃-40℃, other chromatographic conditions are the same as step 2 of this example. The test sample was detected at a detection wavelength of 250nm. The experimental results are as follows Figure 14 As shown in the figure, it can be seen that different column temperatures have slightly different effects on the separation of fingerprint spectra. When the column temperature is 30°C, the chromatographic peak at 15 min is better separated, so the column temperature is preferably 30°C.

[0168] 3.6. Screening of injection volume: Chromatographic conditions: mobile phase B is acetonitrile, mobile phase A is 0.1wt% glacial acetic acid aqueous solution, injection volume is 1μl-3μl, other chromatographic conditions are the same as step 2 of this example. The test sample was detected at a detection wavelength of 250nm. The experimental results are as follows Figure 15 As shown in the figure, it can be seen that different injection volumes have slightly different effects on the separation of fingerprint spectra. When the injection volume is 2 μl, the overall peak shape and peak height are better, so the injection volume is preferably 2 μl.

[0169] 3.7. Flow rate screening: Chromatographic conditions: mobile phase B is acetonitrile, mobile phase A is 0.1wt% glacial acetic acid aqueous solution, flow rate is 0.1ml / min-0.4ml / min, other chromatographic conditions are the same as step 2 of this example. The test sample was detected at a wavelength of 250nm. The experimental results are as follows Figure 16 As shown in the figure, it can be seen that different flow rates have slightly different effects on the separation of fingerprints. When the flow rate is 0.2 ml / min, the most chromatographic peaks are obtained, and the peak shape is better. Therefore, the flow rate is preferably 0.2 ml / min.

[0170] Example 4 Screening of test solution preparation methods

[0171] 1. Extraction method screening: Precisely take 2 ml of Ludaipangshen Oral Liquid (Batch No. 20240123) and place it in a 5 ml volumetric flask. Dilute to the mark with methanol, shake well, centrifuge at 12000 rpm for 10 min, filter the supernatant, and use it as Test Solution 1. Precisely take 2 ml of Ludaipangshen Oral Liquid (Batch No. 20240123) and place it in a 5 ml volumetric flask. Add methanol and ultrasonicate for 5 min, 15 min, and 30 min, respectively, then dilute to the mark, shake well, centrifuge at 12000 rpm for 10 min, filter the supernatant, and use it as Test Solutions 2, 3, and 4.

[0172] The above test solutions were subjected to ultra-high performance liquid chromatography detection. Except that the mobile phase B was acetonitrile and the mobile phase A was 0.1 wt% aqueous glacial acetic acid, the other chromatographic conditions were the same as those in Example 1, and the detection wavelength was 250 nm. The results are shown in Figure 17 From the graph, it can be seen that the extraction time has no obvious effect on peak shape, separation degree, and peak height, so the preparation process of adding methanol and ultrasonication for 5 min is preferred.

[0173] 2. Extraction solvent screening: Precisely take 2 ml of Ludaipangshen Oral Liquid (Batch No. 20240123) and place it in a 5 ml volumetric flask. Add methanol, 50 wt% methanol aqueous solution, ethanol, 50 wt% ethanol aqueous solution, and water, respectively, ultrasonicate for 5 min, dilute to the mark, shake well, centrifuge at 12000 rpm for 10 min, filter the supernatant, and use it as Test Solutions 1, 2, 3, 4, and 5.

[0174] The above test solutions were subjected to ultra-high performance liquid chromatography detection. Except that the mobile phase B was acetonitrile and the mobile phase A was 0.1 wt% aqueous glacial acetic acid, the other chromatographic conditions were the same as those in Example 1, and the detection wavelength was 250 nm. The results are shown in Figure 18 From the graph, it can be seen that when water, 50% methanol, ethanol, and 50% ethanol are used for extraction, ethanol and 50% ethanol have poor separation degree and peak shape at 6-8 min. When methanol, 50% methanol, and water are used for extraction, the peak shape, separation degree, and peak height have no obvious effect, so the extraction solvent can be methanol, 50% methanol, or water.

[0175] 3. Sample amount screening: Precisely take 2 ml, 0.5 ml, 1 ml, and 3 ml of Ludaipangshen Oral Liquid (Batch No. 20240123) and place it in a 5 ml volumetric flask. Add methanol, ultrasonicate for 5 min, dilute to the mark, shake well, centrifuge at 12000 rpm for 10 min, filter the supernatant, and use it as Test Solutions 1, 2, 3, and 4.

[0176] The above test solutions were subjected to ultra-high performance liquid chromatography detection. Except that the mobile phase B was acetonitrile and the mobile phase A was 0.1 wt% aqueous glacial acetic acid, the other chromatographic conditions were the same as those in Example 1, and the detection wavelength was 250 nm. The results are shown in Figure 19 As shown in the figure, when the sampling volume is 2 ml, the peak height is appropriate and the peak separation is good, so the preferred sampling volume is 2 ml.

Claims

1. A method for constructing a fingerprint of Lu Codonopsis pilosula oral liquid, characterized in that The following steps are involved: (1) Using ultra-high performance liquid chromatography to test different batches of test sample solutions to obtain ultra-high performance liquid chromatograms of the test samples; the ultra-high performance liquid chromatography conditions include: Column: ACQUITY Premier HSS T3 column; Mobile phase: Mobile phase A is 0.2 wt% glacial acetic acid in water, and mobile phase B is acetonitrile; The gradient elution program was as follows: from 0 to 11 min, the volume fraction of mobile phase A decreased from 95% to 91%; from 11 to 28 min, the volume fraction of mobile phase A decreased from 91% to 80%; from 28 to 40 min, the volume fraction of mobile phase A decreased from 80% to 71%; from 40 to 43 min, the volume fraction of mobile phase A decreased from 71% to 35%; from 43 to 48 min, the volume fraction of mobile phase A remained unchanged at 35%; (2) The ultra-high performance liquid chromatography (UPLC) chromatogram of the test sample was processed using the Chinese medicine chromatographic fingerprint similarity evaluation system to generate a standard reference fingerprint and determine the common peaks, thus constructing the fingerprint of Ludangshen oral liquid.

2. The construction method according to claim 1, wherein: The ultra-high performance liquid chromatography conditions also include at least one of the following conditions: a. Column temperature: 25-40°C; b. Flow rate: 0.1-0.4 ml / min; c. Injection volume: 1μl-3μl; d. The detector used is a UV-visible spectrophotometer with a detection wavelength of at least one of 220 nm, 250 nm, 268 nm, and 320 nm.

3. The construction method according to claim 1, wherein: The ultra-high performance liquid chromatography conditions also include at least one of the following conditions: a. Column temperature: 30°C; b. Flow rate: 0.2 ml / min; c. Injection volume: 2 μl.

4. The construction method according to any one of claims 1 to 3, characterized in that: The method further comprises the steps of identifying common peaks; selecting adenosine, chlorogenic acid, syringin, geniposide, rutin, codonopsis arginine, codonopsis arginine alcohol, and atractylodes lactone III as reference substances, respectively configuring each reference substance into a reference substance solution with methanol, then mixing each reference substance solution, and fixing the volume with methanol to obtain a mixed reference substance solution; testing the mixed reference substance solution according to the chromatographic conditions of step (1) in claim 1 to obtain a mixed reference substance chromatogram, and identifying the common peaks in the constructed fingerprint of Ludangshen oral liquid according to the mixed reference substance chromatogram, and identifying a total of 8 chromatographic peaks, peak 1 being adenosine, peak 7 being chlorogenic acid, peak 8 being syringin, peak 9 being geniposide, peak 14 being rutin, peak 18 being codonopsis arginine, peak 21 being codonopsis arginine alcohol, and peak 25 being atractylodes lactone III.

5. The construction method according to any one of claims 1 to 3, characterized in that: The preparation method of the test solution is as follows: take 2 ml of Ludangshen oral liquid from different batches, place it in a 5 ml volumetric flask, add solvent and ultrasonicate for 5-30 minutes, adjust the volume to the scale, shake well, centrifuge at 12000 rpm for 10 minutes, take the supernatant and filter to obtain the test solution.

6. A method for detecting the content of quality markers in Ludangshen oral liquid, characterized by: The quality markers are chlorogenic acid, syringin, geniposide, codonopsis pilosula, codonopsis pilosula alcohol and atractylodes lactone III, and the detection method comprises the following steps: (1) Each quality marker is prepared into a solution with methanol, and then the quality marker solutions are mixed and fixed to volume with methanol to obtain a mixed quality marker solution. The mixed quality marker solution is diluted with methanol to a series of different concentrations to obtain a series of reference solution; (2) A series of reference solutions were tested by ultra-high performance liquid chromatography to obtain the peak area of ​​each mass marker, and a standard curve of the concentration and peak area of ​​each mass marker was drawn to obtain the regression equation of each mass marker; (3) The test solution of Ludangshen oral liquid was tested by ultra-high performance liquid chromatography to obtain the peak area of ​​each quality marker, which was substituted into the regression equation to calculate the content of each quality marker; The ultra-high performance liquid chromatography conditions include: Column: ACQUITY Premier HSS T3 column; Mobile phase: Mobile phase A is 0.2 wt% glacial acetic acid in water, and mobile phase B is acetonitrile; The gradient elution program was as follows: from 0 to 11 min, the volume fraction of mobile phase A decreased from 95% to 91%; from 11 to 28 min, the volume fraction of mobile phase A decreased from 91% to 80%; from 28 to 40 min, the volume fraction of mobile phase A decreased from 80% to 71%; from 40 to 43 min, the volume fraction of mobile phase A decreased from 71% to 35%; from 43 to 48 min, the volume fraction of mobile phase A remained unchanged at 35%.

7. The detection method according to claim 6, wherein: Also includes at least one of the following conditions: a. For ultra-high performance liquid chromatography (UPLC) analysis, the column temperature is 25-40°C, the flow rate is 0.1-0.4 ml / min, and the injection volume is 1 μl-3 μl. b. For ultra-high performance liquid chromatography (UPLC), the detection wavelength for chlorogenic acid is 320 nm; the detection wavelength for syringin, codonopsis pilosula, and codonopsis pilosulphol is 268 nm; the detection wavelength for geniposide is 250 nm; and the detection wavelength for atractylodes lactone III is 220 nm. c. Prepare the test solution as follows: Take 2 ml of Lu Codonopsis Oral Liquid and place it in a 5 ml volumetric flask. After adding solvent, sonicate for 5-30 minutes, dilute to the mark, shake well, centrifuge at 12,000 rpm for 10 minutes, and filter the supernatant to obtain the test solution.

8. The fingerprint of Ludangshen oral liquid constructed according to the method for constructing the fingerprint of Ludangshen oral liquid according to any one of claims 1 to 5 is characterized in that: The fingerprint spectrum has a total of 18 common peaks at a detection wavelength of 250nm, namely peak 1, peak 3, peak 5, peak 8, peak 9, peak 10, peak 11, peak 12, peak 14, peak 16, peak 17, peak 18, peak 19, peak 20, peak 21, peak 22, peak 23, and peak 25; at a detection wavelength of 268nm, there are a total of 16 common peaks, namely peak 1, peak 2, peak 3, peak 4, peak 6, peak 8, peak 9, peak 11, peak 13, peak 14, peak 17, peak 18, peak 19, peak 20, peak 21, and peak 24; at a detection wavelength of 220nm, there are a total of 8 common peaks, namely peak 6, peak 8, peak 9, peak 11, peak 13, peak 14, peak 18, and peak 25; at a detection wavelength of 320nm, there are 1 There are 6 common peaks in total, namely peak 3, peak 7, peak 8, peak 13, peak 14 and peak 15; among them, peak 1 is adenosine, peak 7 is chlorogenic acid, peak 8 is syringin, peak 9 is geniposide, peak 14 is rutin, peak 18 is codonopsis pilosula, peak 21 is codonopsis pilosula alcohol, and peak 25 is atractylodes lactone III.

9. The fingerprint according to claim 8 is characterized in that: When the detection wavelength is 250nm, peak 18 codonopsis pilosula glycoside is used as the reference peak, and the relative retention times of the common peaks are: peak 1: 0.0917, peak 3: 0.1679, peak 5: 0.2138, peak 8: 0.4332 , Peak 9: 0.5445 , Peak 10: 0.6032 , Peak 11: 0.6160 , Peak 12: 0.7216 , Peak 14: 0.8258 , Peak 16: 0.8815 , Peak 17: 0.9119 , Peak 18: 1.0000 , Peak 19: 1.0404, Peak 20: 1.1320, Peak 21: 1.1483, Peak 22: 1.1754, Peak 23: 1.2004, Peak 25: 1.3625.

10. The method for constructing the fingerprint of Ludangshen oral liquid according to any one of claims 1 to 5 or / and the method for detecting the content of quality markers of Ludangshen oral liquid according to claim 6 or 7 or / and the use of the fingerprint of Ludangshen oral liquid according to claim 8 or 9 in quality detection and / or quality evaluation and / or quality control of Ludangshen oral liquid.

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