A method for detecting the content of active ingredients in compound licorice oral solution using UPLC

Abstract

The present invention relates to a method for detecting the content of active ingredients in a compound licorice oral solution using UPLC in the field of pharmaceutical analysis. The detection method comprises the following steps: (1) chromatographic conditions: a chromatographic column using octadecylsilane bonded silica gel as a filler; a mobile phase comprising phosphate buffer and acetonitrile; the chromatographic conditions employing mobile phase concentration, gradient elution, and a variable wavelength method; (2) preparation of the phosphate buffer; (3) preparation of a test solution; (4) preparation of a reference solution; and (5) a determination method. The method has the advantages of being able to simultaneously detect the contents of multiple active ingredients, providing intuitive and accurate detection results, a fast detection speed, and readily available raw materials required for detection.

Description

Technical Field

[0001] The invention relates to a method for detecting the content of active ingredients in a compound liquorice oral solution by adopting UPLC method in the field of drug analysis. Technical Background

[0002] Compound Glycyrrhiza Oral Solution, also known as Compound Glycyrrhiza Oral Solution in English, is a brown or brown-black liquid with an aroma and occasional precipitation after long storage. Compound Glycyrrhiza Oral Solution is included in the 2020 edition of the Chinese Pharmacopoeia, Part II. It is mainly composed of licorice fluid extract, compound camphor tincture, guaifenesin, glycerin, concentrated ammonia solution, etc. It is a commonly used medicine for the treatment of coughs and sputum caused by upper respiratory tract infections, bronchitis and colds. Among them, licorice fluid extract is a protective expectorant; compound camphor tincture is an antitussive; guaifenesin is an expectorant and cough suppressant, and has a certain preservative effect; glycerin and concentrated ammonia solution are excipients that can keep the preparation stable and prevent precipitation from forming and settling. The above ingredients constitute a compound preparation with synergistic antitussive and expectorant effects. In the Chinese Pharmacopoeia (2020 edition), the active ingredients of compound licorice oral solution, morphine, guaifenesin and glycyrrhizic acid, were detected by two completely different liquid chromatography systems. When morphine was measured, the sample was first extracted on a solid phase extraction column and then detected on an octylsilane bonded silica gel column. This method is complicated to operate, has low precision, is time-consuming and has high cost. When guaifenesin and glycyrrhizic acid were measured, the requirements for the high performance liquid chromatography system were very high, otherwise the separation degree between guaifenesin and glycyrrhizic acid and their adjacent impurity peaks would not meet the requirements. So far, no method for the simultaneous determination of the content of the active ingredients of compound licorice oral solution has been retrieved. Therefore, the development of a detection method for the simultaneous determination of the active ingredients in compound licorice oral solution has always been a new issue that needs to be solved urgently. Summary of the Invention

[0003] The present invention aims to provide a method for detecting the content of active ingredients in compound liquorice oral solution by UPLC method. The method adopts UPLC method (mobile phase concentration, gradient elution, variable wavelength method) for detection. The method has the advantages of being able to simultaneously detect the contents of multiple active ingredients, readily available reagents, simple operation, short detection cycle, and high precision.

[0004] The object of the present invention is achieved by: a method for detecting the content of active ingredients in compound liquorice oral solution by UPLC method, the detection method comprising the following steps:

[0005] (1) Chromatographic conditions:

[0006] Chromatographic column: Octadecylsilane bonded silica gel as filler

[0007] Mobile phase: phosphate buffer and acetonitrile

[0008] The chromatographic conditions adopt mobile phase concentration, gradient elution, and variable wavelength method;

[0009] (2) Preparation of phosphate buffer

[0010] (3) Preparation of test solution

[0011] (4) Preparation of reference solution

[0012] (5) Determination method:

[0013] Accurately measure the reference solution and the test solution and inject them into the liquid chromatograph to record the chromatogram. In the chromatogram of the test solution, the separation degree between the active ingredient peak and the adjacent chromatographic peak should meet the requirements and be calculated by the peak area according to the external standard method;

[0014] The active ingredient is selected from one or more of morphine, guaifenesin, and glycyrrhizic acid; the specifications of the chromatographic column are selected from (30-150)×(1.8-2.5) mm, 1.6-10 μm; preferably, the specifications of the chromatographic column include (75-150)×(2.0-2.2) mm, 2-3.5 μm; the brand of the chromatographic column is selected from HALO, Waters, Agilent, Dima, and Shiseido; in the chromatogram of the test solution, the theoretical plate number of the morphine peak shall not be less than 2000;

[0015] The preparation method of the phosphate buffer comprises the following steps: taking phosphate, dissolving it in water, adding triethylamine, and adjusting the pH to weak acidity with an organic acid solution; the phosphate is selected from one or more of potassium dihydrogen phosphate and dipotassium hydrogen phosphate; the organic acid is selected from one or more of phosphoric acid and acetic acid; the weak acidity refers to a pH of 3.5-5.5; preferably, the weak acidity refers to a pH of 4-5; the concentration of the organic acid is selected from 1-30%; preferably, the concentration of the organic acid is selected from 5-20%; the mass of the phosphate: the volume of the water: the volume of the triethylamine = (1-10): (500-1500): (0.5-10), the unit of the mass is gram, and the unit of the volume is milliliter; preferably, the mass of the phosphate: the volume of the water: the volume of the triethylamine = (2-5): (800-1250): (1-5), the unit of the mass is gram, and the unit of the volume is milliliter;

[0016] The chromatographic conditions also include: column temperature: 20°C to 45°C; preferably, column temperature: 25°C to 40°C; flow rate: 0.25ml / min to 0.38ml / min; preferably, flow rate: 0.30ml / min to 0.36ml / min; UV detector wavelength: 210-230nm, 240-260nm; preferably, UV detector wavelength: 215-225nm, 245-255nm; injection volume: 0.5μl to 5μl; preferably, injection volume: 0.5μl to 2μl;

[0017] The preparation of the test solution comprises the following steps: taking the compound licorice oral solution, ultrasonicating it, placing it in a volumetric flask, diluting it to the scale with a diluent, shaking it well, filtering it with a microporous filter membrane, and taking the filtrate as the test solution; the ultrasonication time is 1-60 minutes, preferably, the ultrasonication time is 1-30 minutes;

[0018] The preparation of the reference solution comprises the following steps: accurately weighing one or more of morphine, guaifenesin and ammonium glycyrrhizate, placing the mixture in a volumetric flask, adding a diluent to dissolve the mixture and diluting the mixture to the mark, and shaking the mixture to prepare a reference solution; the content of morphine in the reference solution is 0.005-0.03 mg / ml; preferably, the content of morphine in the reference solution is 0.008-0.02 mg / ml; the content of guaifenesin in the reference solution is 0.1-2 mg / ml; preferably, the content of guaifenesin in the reference solution is 0.2-1 mg / ml; the content of glycyrrhizic acid in the reference solution is 0.05-1 mg / ml; preferably, the content of glycyrrhizic acid in the reference solution is 0.15-0.5 mg / ml;

[0019] The preparation of the diluent comprises the following steps: taking acetic acid, water, and methanol and mixing them; the acetic acid is selected from glacial acetic acid; the volume ratio of the acetic acid, water, and methanol is (1-10):(100-1000):(100-1000); preferably, the volume ratio of the acetic acid, water, and methanol is (2-8):(200-800):(200-800); the mobile phase concentration, gradient elution, and variable wavelength method comprise the following steps:

[0020]

[0021] The phosphate buffer and acetonitrile are in a volume ratio;

[0022] Preferably, the mobile phase concentration, gradient elution, and variable wavelength method comprises the following steps:

[0023]

[0024] The phosphate buffer and acetonitrile are in a volume ratio;

[0025] The mobile phase concentration, gradient elution, and variable wavelength method include the following steps:

[0026]

[0027] The phosphate buffer and acetonitrile are in a volume ratio;

[0028] Preferably, the mobile phase concentration, gradient elution, and variable wavelength method comprises the following steps:

[0029]

[0030]

[0031] The phosphate buffer and acetonitrile are in a volume ratio;

[0032] The formula for calculating the peak area by the external standard method is:

[0033]

[0034] A 供 : Peak area of ​​the corresponding active ingredient in the test solution;

[0035] A 对 : Peak area of ​​the corresponding active ingredient in the reference solution;

[0036] C 对 : The concentration of the corresponding active ingredient reference substance in the reference solution, mg / ml;

[0037] P 对 : Purity of the corresponding active ingredient reference substance;

[0038] When calculating the glycyrrhizic acid content, the numerator in the formula should be multiplied by the conversion factor 0.9797.

[0039] The key point of the present invention lies in its detection method, which adopts UPLC (concentration, gradient elution, variable wavelength method) for detection.

[0040] Compared with existing technologies, a method for detecting the content of active ingredients in compound licorice oral solution using UPLC has the advantages of being able to simultaneously detect the contents of multiple active ingredients, readily available reagents, simple operation, shortening the detection time per batch from 2 days to less than 1 day, and high precision. It will be widely used in the field of pharmaceutical analysis. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] The present invention will be described in detail below with reference to the accompanying drawings and embodiments.

[0042] Figure 1 This is the UPLC chromatogram of the reference solution of Example 1 of the present invention.

[0043] Figure 2 This is the UPLC chromatogram of the test solution of Example 1 of the present invention. DETAILED DESCRIPTION

[0044] The following examples will help to understand the present invention, but these embodiments are only for illustrating the present invention and the present invention is not limited to these contents.

[0045] Example 1

[0046] 1. Chromatographic conditions

[0047] Chromatographic column: HALO C18 100×2.1mm 2.7μm, column temperature: 35℃, flow rate: 0.35ml / min, detection wavelength: 220nm, 250nm, injection volume: 1μl.

[0048] 2. Solution Preparation

[0049] 2.1 Mobile phase: Phosphate buffer as mobile phase A, acetonitrile as mobile phase B

[0050] 2.2 Phosphate buffer: Take 3.4 g of potassium dihydrogen phosphate, dissolve it in 1000 ml of water, add 2.5 ml of triethylamine, and adjust the pH to 4.5 with 10% phosphoric acid solution.

[0051] 2.3 Diluent: Take 5 ml of glacial acetic acid, add water to 500 ml, mix well, add 500 ml of methanol, shake well, and you have it.

[0052] 2.4 Test solution: Take 1 bottle of compound licorice oral solution (Northeast Pharmaceutical Group Shenyang No. 1 Pharmaceutical Co., Ltd., 100 ml / bottle, batch number 191206), ultrasonicate for 10 minutes, let cool, accurately measure 5 ml of compound licorice oral solution, place it in a 50 ml volumetric flask, dilute to the scale with diluent, shake well, filter with a 0.22 μm filter membrane, and take the filtrate as the test solution.

[0053] 2.5 Reference solution: Accurately weigh 10 mg of morphine reference substance, place it in a 100 ml volumetric flask, dissolve it with diluent and dilute it to the mark, shake well, and use it as the morphine reference substance stock solution; accurately weigh 25 mg of guaifenesin reference substance and 12.5 mg of ammonium glycyrrhizate reference substance, place them in the same 50 ml volumetric flask, accurately add 5 ml of morphine reference substance stock solution, dissolve it with diluent and dilute it to the mark, shake well, and use it as the reference substance solution.

[0054] 3. Determination method

[0055] The mobile phase was phosphate buffer and acetonitrile, the concentration was gradient, and the wavelength was varied;

[0056]

[0057] In the above table, the ratio of mobile phase A to mobile phase B is by volume.

[0058] See attached Figure 2 In the chromatogram of the test solution, the theoretical plate number of the morphine peak is 3172, and the morphine peak and the adjacent chromatographic peak ( Figure 2 The separation degree of the chromatographic peaks (the adjacent chromatographic peaks indicated by the arrow closest to the morphine peak) is 1.8, and the separation degree of the guaifenesin peak and the adjacent chromatographic peaks ( Figure 2 The separation degree of glycyrrhizic acid and the adjacent chromatographic peak (the arrow closest to the guaifenesin peak) is 1.8, and the separation degree of glycyrrhizic acid and the adjacent chromatographic peak ( Figure 2 The resolution of the adjacent chromatographic peaks (indicated by the arrow closest to the glycyrrhizic acid peak) is 1.6.

[0059] Accurately measure 0.5μl to 5μl of the reference solution and the test solution and inject them into the liquid chromatograph to record the chromatogram.

[0060] 4. Calculation results

[0061] 4.1 Morphine content

[0062] Reference solution 1 morphine peak area: 93.78 95.00 93.80 94.68 94.04

[0063] Mean: 94.26 RSD: 0.6%

[0064] Reference solution 2 Morphine Peak Area: 95.76 96.45 Average: 96.10

[0065] Average sample weight of reference substance: (10.49+10.81) / 2=10.65

[0066] Average peak area of ​​reference substance: (94.26+96.10) / 2=95.18

[0067] Test solution 1 Morphine Peak Area: 96.48 97.39 Average: 96.93

[0068] Test solution 2 Morphine Peak Area: 96.42 96.74 Average: 96.58

[0069]

[0070]

[0071] Average value: 0.1013 mg / ml

[0072] 4.2 Guaifenesin content

[0073] Reference solution 1 guaifenesin peak area: 130.46 130.87 130.92 130.93 130.43

[0074] Mean: 130.72 RSD: 0.2%

[0075] Reference solution 2 Guaifenesin peak area: 128.57 128.87 Average: 128.72

[0076] Average sample weight of reference substance: (25.37+25.13) / 2=25.25

[0077] Average peak area of ​​reference substance: (130.72+128.72) / 2=129.72

[0078] Test solution 1 Guaifenesin peak area: 126.36 127.13 Average: 126.74

[0079] Test solution 2 Guaifenesin peak area: 126.16 126.28 Average: 126.22

[0080]

[0081]

[0082] Average: 4.904 mg / ml

[0083] 4.3 Glycyrrhizic acid content

[0084] Reference solution 1 glycyrrhizic acid peak area: 523.40 535.58 527.67 527.54 523.36

[0085] Average: 527.51 RSD: 1.0%

[0086] Reference solution 2 Glycyrrhizic acid peak area: 523.74 525.28 Average: 524.51

[0087] Average sample weight of reference substance: (12.64+12.70) / 2=12.67

[0088] Average peak area of ​​reference substance: (527.51+524.51) / 2=526.01

[0089] Test solution 1 Glycyrrhizic acid peak area: 587.27 589.43 Average: 588.35

[0090] Test solution 2 Glycyrrhizic acid peak area: 584.91 585.81 Average: 585.36

[0091]

[0092]

[0093] Average: 2.61 mg / ml

[0094] Table 1 Determination results of active ingredients in compound licorice oral solution

[0095]

[0096] Example 2

[0097] Methodological investigation

[0098] This study systematically investigated the linearity, repeatability, accuracy and stability of this method.

[0099] 1. Linearity test

[0100] 1.1 Morphine linearity test

[0101] Accurately weigh 10 mg of the morphine reference substance and place it in a 100 ml volumetric flask. Add diluent to dissolve and dilute to the scale, and shake well. Then accurately measure 4 ml, 4.5 ml, 5 ml, 5.5 ml, and 6 ml and place them in five 50 ml volumetric flasks, dilute to the scale with diluent, and shake well to obtain a series of reference substance solutions with concentrations of 80%, 90%, 100%, 110%, and 120%. The samples were injected and analyzed according to the chromatographic conditions in Example 1. A standard curve was plotted with morphine concentration (X) as the abscissa and peak area (Y) as the ordinate. Linear regression was performed. The results showed that the morphine concentration had a good linear relationship in the range of 7.6602 μg / ml to 11.4903 μg / ml.

[0102] 1.2 Guaifenesin linearity test

[0103] Accurately weigh 10 mg, 11.2 mg, 12.5 mg, 13.8 mg, and 15 mg of the guaifenesin reference substance and place them in 25 ml volumetric flasks, dissolve them in diluent, dilute to the mark, and shake well to obtain a series of reference substance solutions with concentrations of 80%, 90%, 100%, 110%, and 120%. Samples were injected and analyzed according to the chromatographic conditions in Example 1. A standard curve was plotted with guaifenesin concentration (X) as the abscissa and peak area (Y) as the ordinate. Linear regression was performed. The results showed that the guaifenesin concentration had a good linear relationship in the range of 402.78 μg / ml to 635.84 μg / ml.

[0104] 1.3 Glycyrrhizic acid linearity test

[0105] Accurately weigh 10.8 mg, 12.1 mg, 13.5 mg (equivalent to 12.5 mg of glycyrrhizic acid), 14.8 mg, and 16.2 mg of ammonium glycyrrhizate reference substance, respectively, and place them in 50 ml measuring flasks, dissolve them in diluent and dilute to the scale, shake well, and obtain a series of reference substance solutions with concentrations of 80%, 90%, 100%, 110%, and 120%. Sample injection analysis was performed according to the chromatographic conditions in Example 1, with glycyrrhizic acid concentration (X) as the abscissa and peak area (Y) as the ordinate, a standard curve was plotted, and linear regression was performed. The results showed that the linear relationship was good in the range of glycyrrhizic acid concentration of 194.95 μg / ml to 310.19 μg / ml.

[0106] Table 2 Results of linearity test for active ingredients in compound licorice oral solution

[0107] Active ingredient name Linear equations Correlation coefficient morphine Y=9.61328X-0.96744 r＝0.99994 Guaifenesin Y=0.2516X+5.0451 r＝0.9991 Glycyrrhizic acid Y=2.1203X+46.4767 r＝0.9990

[0108] 2. Repeatability test

[0109] Take one bottle of this product, sonicate for 10 minutes, let cool, and accurately measure six 5ml portions of this product. Place each into a 50ml volumetric flask, dilute to the mark with diluent, shake well, filter through a 0.22μm filter, and take the filtrate as the test solution. Sample analysis was performed according to the chromatographic conditions in Example 1. Based on the measured content results (see Table 3), the calculated RSD values ​​for morphine were 0.2%, guaifenesin 0.4%, and glycyrrhizic acid 0.6%, indicating good reproducibility of the method.

[0110] Table 3 Repeatability test results of compound licorice oral solution

[0111]

[0112] 3. Accuracy test

[0113] Blank excipient solution: Weigh the prescribed amount of excipient, place it in a 100ml volumetric flask, add water to dissolve and dilute to the scale, shake well, and use it as the blank excipient solution.

[0114] Accurately weigh 10 mg of the morphine reference substance and place it into a 100 ml volumetric flask. Dissolve the solution in diluent and dilute to the mark. Shake well to prepare the morphine reference substance stock solution. Accurately weigh three portions of 20 mg, 25 mg, and 30 mg of the guaifenesin reference substance and 10 mg, 12.5 mg, and 15 mg of the ammonium glycyrrhizate reference substance. Place the solution into nine 50 ml volumetric flasks. Accurately add three portions of 4 ml, 5 ml, and 6 ml of the morphine reference substance stock solution. Add 5 ml of the blank excipient solution to each flask. Dissolve the solution in diluent and dilute to the mark. Shake well. Filter through a 0.22 μm filter membrane. The filtrate is used as the test solution. Sample injection and analysis were performed according to the chromatographic conditions in Example 1, and the recovery was calculated according to the formula "recovery = recovered amount / added amount × 100%" (see Table 4). The results showed that the average recovery of morphine was 99.5% with an RSD value of 0.7%; the average recovery of guaifenesin was 99.6% with an RSD value of 0.7%; and the average recovery of glycyrrhizic acid was 100.0% with an RSD value of 0.5%. The RSDs were all less than 1%, indicating that the sample recovery was good.

[0115] Table 4 Accuracy determination results of compound licorice oral solution

[0116]

[0117]

[0118]

[0119] 4. Stability test

[0120] According to the chromatographic conditions in Example 1, the samples were injected at 0 h, 1 h, 2 h, 4 h, 6 h, 8 h, 12 h, and 18 h, and the corresponding contents were calculated (see Table 5). The RSD value of morphine was 0.4%, the RSD value of guaifenesin was 0.4%, and the RSD value of glycyrrhizic acid was 0.4%, indicating that the sample had good stability within 18 hours.

[0121] Table 5 Stability test results of compound licorice oral solution

[0122]

Claims

1. A method for detecting the content of active ingredients in compound liquorice oral solution using UPLC, characterized in that: The detection method comprises the following steps: (1) Chromatographic conditions: Chromatographic column: Octadecylsilane bonded silica gel as filler Mobile phase: phosphate buffer and acetonitrile The chromatographic conditions adopt mobile phase concentration, gradient elution, and variable wavelength method; (2) Preparation of phosphate buffer (3) Preparation of test solution (4) Preparation of reference solution (5) Determination method: Accurately measure the reference solution and the test solution and inject them into the liquid chromatograph to record the chromatogram. In the chromatogram of the test solution, the separation degree between the active ingredient peak and the adjacent chromatographic peak should meet the requirements and be calculated by the peak area according to the external standard method; The active ingredient is selected from morphine, guaifenesin and glycyrrhizic acid; The preparation method of the phosphate buffer comprises the following steps: taking phosphate, dissolving it in water, adding triethylamine, and adjusting the pH to weak acidity with an organic acid solution; the weak acidity refers to a pH of 3.5-5.5; The mobile phase concentration, gradient elution, and variable wavelength method include the following steps: The phosphate buffer and acetonitrile are in a volume ratio.

2. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 1, characterized in that: The specifications of the chromatographic column are selected from (30-150)×(1.8-2.5) mm, 1.6-10 μm; the brand of the chromatographic column is selected from HALO, Waters, Agilent, Dima, and Shiseido; in the chromatogram of the test solution, the theoretical plate number of the morphine peak shall not be less than 2000.

3. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 2, characterized in that: The specifications of the chromatographic column are selected from (75-150) × (2.0-2.2) mm, 2-3.5 μm.

4. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 1, characterized in that: The phosphate is selected from one or more of potassium dihydrogen phosphate and dipotassium hydrogen phosphate; the organic acid is selected from one or more of phosphoric acid and acetic acid; the concentration of the organic acid is selected from 1-30%; the mass of the phosphate: the volume of the water: the volume of the triethylamine = 1-10:500-1500:0.5-10, the unit of the mass is gram, and the unit of the volume is milliliter.

5. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 4, characterized in that: The weak acidity refers to a pH of 4-5; the concentration of the organic acid is selected from 5-20%; the mass of the phosphate: the volume of the water: the volume of the triethylamine = 2-5:800-1250:1-5, the unit of the mass is gram, and the unit of the volume is milliliter.

6. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 1, characterized in that: The chromatographic conditions also include: column temperature: 20°C to 45°C; flow rate: 0.25ml / min to 0.38ml / min; injection volume: 0.5μl to 5μl.

7. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 6, characterized in that: Column temperature: 25℃~40℃; flow rate: 0.30ml / min~0.36ml / min; injection volume: 0.5μl~2μl.

8. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 1, characterized in that: The preparation of the test solution comprises the following steps: taking the compound licorice oral solution, ultrasonicating it, placing it in a volumetric flask, diluting it to the scale with a diluent, shaking it evenly, filtering it with a microporous filter membrane, and taking the filtrate as the test solution; the ultrasonication time is 1-60 minutes.

9. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 8, characterized in that: The ultrasonic time is 1-30 min.

10. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 1, characterized in that: The preparation of the reference solution comprises the following steps: accurately weighing one or more of morphine, guaifenesin, and ammonium glycyrrhizate, placing the mixture in a volumetric flask, adding a diluent to dissolve the mixture and diluting the mixture to the mark, and shaking the mixture to prepare a reference solution; the content of morphine in the reference solution is 0.005-0.03 mg / ml; the content of guaifenesin in the reference solution is 0.1-2 mg / ml; and the content of glycyrrhizic acid in the reference solution is 0.05-1 mg / ml.

11. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 10, characterized in that: The content of morphine in the reference solution is 0.008-0.02 mg / ml; the content of guaifenesin in the reference solution is 0.2-1 mg / ml; and the content of glycyrrhizic acid in the reference solution is 0.15-0.5 mg / ml.

12. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC method according to claim 8 or 10, characterized in that: The preparation of the diluent comprises the following steps: taking acetic acid, water and methanol and mixing them evenly; the acetic acid is selected from glacial acetic acid; and the volume ratio of the acetic acid, water and methanol is 1-10:100-1000:100-1000.

13. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 12, characterized in that: The volume ratio of the acetic acid, water and methanol is 2-8:200-800:200-800.

14. The method for detecting the content of active ingredients in compound liquorice oral solution by UPLC according to claim 1, characterized in that: The formula for calculating the peak area by the external standard method is: ， A 供 : Peak area of ​​the corresponding active ingredient in the test solution; A 对 : Peak area of ​​the corresponding active ingredient in the reference solution; C 对 : The concentration of the corresponding active ingredient reference substance in the reference solution, mg / ml; P 对 : Purity of the corresponding active ingredient reference substance; When calculating the glycyrrhizic acid content, the numerator in the formula should be multiplied by the conversion factor 0.9797.

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