Method for determining contents of multiple components in Sanstrocao-Feilong mixture and application of Sanstrocao-Feilong mixture

Abstract

The invention discloses a method for determining the content of multiple components in a Sapococcum / Sapococcum mixture and application. The method comprises the steps of preparation of a test solution, preparation of a mixed reference solution, determination of chromatographic conditions, content determination and the like. According to the method for determining the content of the multiple components in the Sanyone mixture, the content of the eight components in the Sanyone mixture can be determined at the same time within 50 min through optimization of chromatographic conditions, the analysis and detection time is greatly shortened, waste of reagent testing reagents is avoided, energy consumption of equipment is reduced, and meanwhile, the detection efficiency is improved. According to the present invention, the multi-index components are determined, such that the overall quality condition of the preparation can be efficiently and comprehensively reflected, such that the drug quality control standard is substantially improved, the medication safety of the patient can be easily ensured, the drug quality can be easily improved, and the important value is provided for the quality control and improvement and the clinical promotion and application of the Sapococcum / Sapococcum mixture.

Description

Technical Field

[0001] The invention belongs to the field of traditional Chinese medicine analysis and detection, and specifically relates to a method for determining the contents of multiple components in a Sanao Yulong mixture and an application thereof. Background Art

[0002] Sanao Yulong Mixture is made of 8 kinds of Chinese medicines, including dried tangerine peel, licorice, fried bitter almonds, dried houttuynia, ephedra, earthworms, fried yam, and Fritillaria thunbergii. The approval number is: Wanyaozhizi Z20210005000. Sanao Yulong Mixture has the effects of clearing the lungs and relieving exterior symptoms, benefiting the lungs and relieving asthma, relieving cough and eliminating phlegm. It has a history of more than 30 years in clinical practice, and its efficacy has been widely recognized by patients.

[0003] Chinese medicine mixture is a compound dosage form containing multiple components extracted from Chinese medicine pieces with water or other solvents through appropriate processes. Sanao Yulong mixture has the disadvantages of being inconvenient to carry and take, a large single dose, and poor patient compliance. Therefore, the quality control of Sanao Yulong mixture and the development and utilization of preparations need to be solved urgently. The document "HPLC method for determining the content of hesperidin in Sanao Yulong mixture" established a method for determining the content of hesperidin in Sanao Yulong mixture. It only determined the content of hesperidin as a component and could not reflect the overall quality. However, Chinese medicine compound has the characteristics of multiple components, multiple targets, and comprehensive effects. The composition of effective substances is complex, and the efficacy of Chinese medicine compound is not determined by a single or high-content component, but is the result of the synergistic effect of multiple components. Therefore, in order to ensure the quality of Chinese medicine compound and the clinical efficacy, it is necessary to determine the content of multiple components. Summary of the invention

[0004] In view of the deficiencies in the prior art, the object of the present invention is to provide a method for determining the contents of multiple components in Sanao Yulong mixture. This method can simultaneously determine the contents of eight components in Sanao Yulong mixture, including amygdalin, liquiritin, hyperoside, ferulic acid, naringin, hesperidin, quercetin, and ammonium glycyrrhizinate, thereby greatly shortening the analysis and detection time, and having high accuracy and good repeatability, thereby greatly improving the quality control standards of drugs.

[0005] The present invention is achieved through the following technical solutions:

[0006] A method for determining the contents of multiple components in Sanao Yulong mixture comprises the following steps:

[0007] (a) Preparation of the test solution: accurately pipette the Sanaoyulong mixture, dilute it with methanol, filter it, and take the filtrate to obtain the test solution;

[0008] (b) Preparation of mixed reference solution: accurately weigh hesperidin, rutin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin reference substances, dilute to volume with methanol, filter, and take the filtrate to obtain a mixed reference solution;

[0009] (c) Determination of chromatographic conditions: Chromatographic column: octadecylsilane bonded silica gel as filler; mobile phase: acetonitrile (A)-0.1% formic acid solution (B); gradient elution program: 0-10 min, 10% A→15% A; 10-24 min, 15% A→25% A; 24-28 min, 25% A→26% A; 28-30 min, 26% A→30% A; 30-40 min, 30% A→70% A; 40-45 min, 70% A→10% A; 45-50 min, 10% A; dual wavelength method of 254 nm and 283 nm was used for detection, 254 nm was used as the detection wavelength for amygdalin, hyperoside, quercetin, and ammonium glycyrrhizinate, and 283 nm was used as the detection wavelength for liquiritin, ferulic acid, hesperidin, and naringin;

[0010] (d) Content determination: The mixed reference solution and the test solution were respectively injected into the HPLC, and the determination was carried out according to the above chromatographic conditions. The regression equation of each reference substance was established, and the contents of hesperidin, rutin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin in the test solution were calculated by the external standard method.

[0011] Preferably, in step (a), the mass concentration of methanol is 100%.

[0012] Preferably, in step (a), the volume ratio of the Sanaoyulong mixture to methanol is 1:3-5, more preferably 1:4.

[0013] Preferably, in step (b), the mass concentration of methanol is 75% to 85%.

[0014] Preferably, in step (c), the chromatographic column is East Laboratory Epic C18, with specifications of 4.6 mm×250 mm and 5 μm.

[0015] Preferably, in step (c), the flow rate is 0.6-1.0 mL / min; the column temperature is 25-35° C.; and the injection volume is 10 μL.

[0016] Preferably, in step (d),

[0017] The regression equation of hesperidin is Y=7050.4X+26407, and the correlation coefficient R 2 is 0.9991;

[0018] The regression equation of naringenin is Y=20156X+3093.5, and the correlation coefficient R 2 is 1;

[0019] The regression equation of ammonium glycyrrhizinate is Y=9163.9X-22574, and the correlation coefficient R 2 is 0.9994;

[0020] The regression equation of liquiritin is Y=19811X-587.2, and the correlation coefficient R 2 is 1;

[0021] The regression equation of ferulic acid is Y=40351X+95.8, and the correlation coefficient R 2 is 1;

[0022] The regression equation of amygdalin is Y=365.2X-2461.7, and the correlation coefficient R 2 is 0.9997;

[0023] The hyperoside regression equation is Y=38283X+358.54, and the correlation coefficient R 2 is 1;

[0024] The regression equation of quercetin is Y=40127X+11836, and the correlation coefficient R 2 is 0.9993.

[0025] The present invention also provides the application of the method for determining the contents of multiple components in the Sanao Yulong mixture in the quality control of the Sanao Yulong mixture.

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] The present invention establishes a method for determining the contents of multiple components in Sanao Yulong mixture. Through the optimization of chromatographic conditions, the method can simultaneously determine the contents of 8 components of Sanao Yulong mixture within 50 minutes, which greatly shortens the analysis and detection time, avoids the waste of test reagents, and reduces the energy consumption of equipment. At the same time, since the method determines multiple index components, it can efficiently and comprehensively reflect the overall quality of the preparation, thereby greatly improving the quality control standards of drugs, which is beneficial to ensuring the safety of patients' medication and improving the quality of drugs, and is of great value to the quality control and improvement of Sanao Yulong mixture and its clinical promotion and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 It is a chromatogram of a specific test at 254nm and 283nm detection wavelengths, wherein: 1- amygdalin; 2- glycyrrhizin; 3- hyperoside; 4- ferulic acid; 5- naringin; 6- hesperidin; 7- quercetin; 8- ammonium glycyrrhizinate;

[0029] Figure 2 It is the chromatogram of the test sample using different detection wavelengths;

[0030] Figure 3 It is the chromatogram of the test sample using different mobile phases at a detection wavelength of 254nm;

[0031] Figure 4 It is the chromatogram of the test sample using different mobile phases at a detection wavelength of 283nm;

[0032] Figure 5 It is the chromatogram of the test sample at different column temperatures at a detection wavelength of 254 nm;

[0033] Figure 6 It is the chromatogram of the test sample at different column temperatures at a detection wavelength of 283 nm;

[0034] Figure 7 It is the chromatogram of the test product at different flow rates at a detection wavelength of 254nm;

[0035] Figure 8 It is the chromatogram of the test product at different flow rates at a detection wavelength of 283nm;

[0036] Fig. 9 It is the chromatogram of the test sample using different dilution solvents at a detection wavelength of 283nm;

[0037] Fig.10 It is the chromatogram of the test sample using different gradient elution programs at a detection wavelength of 283nm. DETAILED DESCRIPTION

[0038] In order to make those skilled in the art better understand the technical solution of the present invention, the technical solution of the present invention is further described in detail below in conjunction with the accompanying drawings and specific implementation methods. However, the implementation methods of the present invention are not limited to the following embodiments.

[0039] 1. Instruments and reagents

[0040] 1.1 Instrument

[0041] Waters ACQITY Arc HPLC instrument was purchased from Waters Corporation of the United States; YL-060ST ultrasonic cleaner was purchased from Shenzhen Yulu Cleaning Equipment Co., Ltd.; AB135-S electronic analytical balance was purchased from Sartorius Scientific Instruments (Beijing) Co., Ltd.

[0042] 1.2 Drug testing

[0043] Reference substances of hesperidin, naringin, ammonium glycyrrhizinate, amygdalin, ferulic acid, hyperoside and quercetin (batch numbers PS011588, PS011583, PS011584, PS011718, PS012244, PS011442 and PS012980, with purity ≥98.0%) were purchased from Chengdu Pusi Biotechnology Co., Ltd.; glycyrrhizin (batch number 551-15-5, purity ≥98.0%) was purchased from Beijing Zhongke Quality Inspection Biotechnology Co., Ltd.

[0044] 15 batches of Sanao Yulong mixture (batch numbers are 24013001, 24012601, 24012301, 24030601, 24010801, 24012901, 24012801, 24012701, 24031901, 24031201, 24032801, 24032701, 24032901, 24030801, 24030701, numbered S1 to S15) were all provided by Taihe County Hospital of Traditional Chinese Medicine; acetonitrile and methanol were chromatographically pure, formic acid was analytically pure, and water was Wahaha purified water.

[0045] 2 Methods and Results

[0046] 2.1 Chromatographic conditions

[0047] The chromatographic column was East Laboratory Epic C18 (4.6 mm × 250 mm, 5 μm); the mobile phase was acetonitrile (A)-0.1% formic acid solution (B), with gradient elution (0-10 min, 10% A→15% A; 10-24 min, 15% A→25% A; 24-28 min, 25% A→26% A; 28-30 min, 26% A→30% A; 30-40 min, 30% A→70% A; 40-45 min, 70% A→10% A; 45-50 min, 10% A); the flow rate was 0.8 mL / min; the column temperature was 30°C; the injection volume was 10 μL; the detection wavelengths were 254 nm (amygdalin, hyperoside, quercetin, ammonium glycyrrhizinate) and 283 nm (liquiritigenin, ferulic acid, hesperidin, naringin).

[0048] 2.2 Solution preparation

[0049] 2.2.1 Preparation of test solution: Accurately pipette Sanao Yulong into a volumetric flask, add methanol to the mark, shake evenly, filter through an organic microfiltration membrane, and take the filtrate to prepare the Sanao Yulong mixture test solution.

[0050] 2.2.2 Preparation of mixed reference solution: Accurately weigh hesperidin, rutin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin reference substances, and use 80% methanol as solvent to prepare mixed reference solutions with mass concentrations of 300.00, 66.40, 74.80, 42.00, 12.00, 100.00, 12.40 and 15.70 μg / mL, respectively.

[0051] 2.3 Investigation of linear relationship

[0052] The mixed reference solution prepared in "2.2.2" was diluted by 1, 2, 4, 8, and 16 times to prepare solutions of various concentrations. The solutions were measured according to the chromatographic conditions in "2.1". The standard curve was drawn with the mass concentration of the solution as the abscissa (x) and the peak area as the ordinate (Y). The results showed that within a certain concentration range, the eight components had good correlation, as shown in Table 1.

[0053] Table 1 Linear relationship investigation

[0054]

[0055] 2.4 Methodological Review

[0056] 2.4.1 Precision test

[0057] Accurately pipette the same mixed reference solution and inject it 6 times according to the method under "2.1". The RSDs of the peak areas of hesperidin, naringin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin were calculated to be 0.02%, 0.43%, 0.54%, 0.40%, 0.53%, 0.74%, 0.41% and 0.41%, respectively, indicating that the instrument has good precision.

[0058] 2.4.2 Stability test

[0059] A group of samples of Sanao Yulong mixture were randomly selected to prepare test solution. The samples were injected and measured at 0, 2, 4, 8, 12 and 24 h according to the chromatographic conditions under "2.2". The RSDs of the peak areas of hesperidin, naringin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin were calculated to be 0.44%, 0.63%, 1.21%, 0.79%, 0.85%, 0.91%, 0.81% and 0.61%, respectively, indicating that the test solution had good stability within 24 h.

[0060] 2.4.3 Repeatability test

[0061] Six test solutions were prepared in parallel from the same group of Sanaoyulong mixture samples and determined according to the chromatographic conditions under "2.2". The RSDs of the peak areas of hesperidin, naringin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin were calculated to be 0.36%, 0.90%, 1.20%, 1.09%, 0.71%, 1.11%, 1.61% and 0.61%, respectively, indicating that the method has good repeatability.

[0062] 2.4.4 Sample recovery test

[0063] Using methanol as solvent, prepare mixed reference solutions with peak mass concentrations of 0.541, 0.138, 0.231, 0.063, 0.006, 0.310, 0.006, and 0.011 mg / mL for hesperidin, naringin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside, and quercetin, respectively; accurately pipette 0.5 mL each of the mixed reference solution, pure water, and Sanao Yulong mixture test solution of known concentration into a 5 mL volumetric flask, dilute to the mark with methanol, and prepare 6 replicates in parallel; then determine them separately according to the chromatographic conditions under "2.2". The calculated average recoveries of the above eight components were 99.00%, 103.31%, 102.27%, 99.29%, 99.05%, 101.08%, 97.1% and 103.42%, respectively; the RSDs were 0.47%, 0.75%, 1.98%, 1.13%, 0.83%, 2.07%, 1.84% and 0.78% (n=6), respectively. The results are shown in Table 2.

[0064] Table 2 Sample recovery test results

[0065]

[0066]

[0067]

[0068] 2.6 Sample content determination

[0069] Fifteen batches of Sanao Yulong mixture were taken to prepare test solutions, and the samples were injected and measured according to the chromatographic conditions under "2.2". The average contents of hesperidin, naringin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin were calculated to be 517.06, 120.89, 195.66, 57.06, 296.04, 6.75, 8.89 and 5.93 μg / mL, respectively. The results are shown in Table 3.

[0070] Table 3 Content determination results of 8 components in 15 batches of Sanaoyulong mixture

[0071]

[0072] 3 Optimization of chromatographic conditions

[0073] 3.1 Wavelength Optimization

[0074] The present invention investigates 8 different detection wavelengths of 220nm, 240nm, 254nm, 260nm, 283nm, 300nm, 320nm, and 340nm. Figure 1 As shown in the figure, combined with the maximum absorption wavelength of each component and the actual measurement results, the dual wavelength method of 254 and 283 nm was finally used for detection. 254 nm was selected as the detection wavelength of four components, including amygdalin, hyperoside, quercetin, and ammonium glycyrrhizinate, and 283 nm was selected as the detection wavelength of four components, including liquiritin, ferulic acid, hesperidin, and naringin. The chromatograms of the test solution at each wavelength are shown in Figure 2 .

[0075] 3.2 Optimization of mobile phase

[0076] The present invention investigates four different mobile phase systems: methanol (A)-water (B), methanol (A)-0.1% formic acid (B), acetonitrile (A)-water (B), and acetonitrile (A)-0.1% formic acid (B). Figure 3-4 As shown in the figure, since the elution ability of the methanol-water mobile phase system is weaker than that of the acetonitrile-water mobile phase system, the chromatographic peak is relatively late, which increases the analysis time, so the acetonitrile-water mobile phase system with stronger elution ability is selected. By comparing the acetonitrile-water and acetonitrile 0.1% formic acid mobile phase systems, it can be found that after adding 0.1% formic acid to the water phase, the number of chromatographic peaks and the response are increased, so acetonitrile (A)-0.1% formic acid (B) is finally selected as the mobile phase.

[0077] 3.3 Optimization of column temperature

[0078] The present invention investigated three different column temperatures of 25°C, 30°C, and 35°C. Figure 5-6 As shown, when the column temperature is 30°C, the peak shapes of the chromatograms are relatively symmetrical and the separation is good, so 30°C is finally selected as the column temperature of the chromatographic column of the present invention;

[0079] 3.4 Optimization of flow rate

[0080] The present invention investigated three different flow rates of 0.6 mL / min, 0.8 mL / min, and 1.0 mL / min. Figure 7-8 As shown in the figure, with the increase of flow rate, the sample peak time is also advanced, but when the flow rate is 0.8mL / min, the separation of chromatographic peaks is the best and the solvent is saved, so the flow rate is finally selected as 0.8mL / min.

[0081] 3.5 Optimization of test sample dilution ratio

[0082] The present invention investigates the dilution of the test sample: methanol solvent at four different ratios of 4:1, 1:1, 1:4, and 1:9. Fig. 9 As shown in the figure, when the ratio of the test sample to methanol solvent was 4:1 and 1:1, the peak at the retention time of 14.5min was bifurcated and the filtration was difficult. When the ratio of the test sample to methanol solvent was 1:9, there were fewer chromatographic peaks, many chemical components did not respond, and the separation was low. Therefore, the final ratio of the test sample to methanol solvent was 1:4, and the chromatogram peak shape was more symmetrical.

[0083] 3.6 Optimization of gradient elution program

[0084] The present invention investigates 6 different gradient elution programs. Since the chromatographic peak information is richer at a detection wavelength of 283 nm and all 8 components to be measured respond, the chromatogram at a detection wavelength of 283 nm is used as an example to illustrate the optimization process of the gradient elution program.

[0085] Method 1: acetonitrile (A)-0.1% formic acid solution (B), 0-15 min, 10% A→30% A; 15-19 min, 30% A→31% A; 19-29 min, 31% A→33% A; 29-35 min, 33% A→40% A; 35-45 min, 40% A→70% A; 45-50 min, 70% A→10% A; 50-55 min, 10% A.

[0086] Method 2: acetonitrile (A)-0.1% formic acid solution (B), 0-8 min, 10% A→15% A; 8-20 min, 15% A→25% A; 20-28 min, 25% A→26% A; 28-32 min, 26% A→30% A; 32-40 min, 30% A→70% A; 40-45 min, 70% A→10% A; 45-50 min, 10% A.

[0087] Method 3: acetonitrile (A)-0.1% formic acid solution (B), 0-10 min, 10% A→15% A; 10-24 min, 15% A→25% A; 24-28 min, 25% A→26% A; 28-30 min, 26% A→30% A; 30-40 min, 30% A→40% A; 40-50 min, 40% A→70% A; 50-55 min, 60% A→10% A; 55-60 min, 10% A.

[0088] Method 4: acetonitrile (A)-0.1% formic acid solution (B), 0-10 min, 10% A→15% A; 10-24 min, 15% A→25% A; 24-28 min, 25% A→26% A; 28-30 min, 26% A→30% A; 30-45 min, 30% A→33% A; 45-50 min, 33% A→40% A; 50-55 min, 40% A→60% A, 55-60 min, 60% A→10% A; 60-65 min, 10% A.

[0089] Method 5: acetonitrile (A)-0.1% formic acid solution (B), 0-10 min, 10% A→15% A; 10-30 min, 15% A→30% A; 30-40 min, 30% A→60% A; 40-45 min, 60% A→10% A; 45-50 min, 10% A.

[0090] Method 6: acetonitrile (A)-0.1% formic acid solution (B), 0-10 min, 10% A→15% A; 10-24 min, 15% A→25% A; 24-28 min, 25% A→26% A; 28-30 min, 26% A→30% A; 30-40 min, 30% A→70% A; 40-45 min, 70% A→10% A; 45-50 min, 10% A.

[0091] like Fig.10 As shown, method 1 has too high initial acetonitrile concentration in the mobile phase, and each component peaks within 19 minutes, and the component signal peaks are piled up and the number is small, which is not suitable as an analysis condition; method 2 has fewer chromatographic peaks, many chemical components have no response, and the separation degree is low; method 3 is improved based on method 2, and it can be seen that the chromatographic peaks increase, but the signal peaks are too concentrated, and deformed peaks appear at retention times of 13.5 minutes and 18.5 minutes; method 4 is further optimized than method 3, but the separation degree is poor at the retention time of 35 minutes; methods 5 and 6 are further optimized than method 4, and the chromatographic peak separation effect of gradient 5 is greatly improved, but the separation of individual peaks is still poor; and each chromatographic peak in the fingerprint spectrum obtained by method 6 has achieved a good separation effect, so method 6 is finally selected as the gradient elution procedure of the present invention.

Claims

1. A method for determining the contents of multiple components in Sanao Yulong mixture, characterized in that: The steps include: (a) Preparation of the test solution: accurately pipette the Sanaoyulong mixture, dilute it with methanol, filter it, and take the filtrate to obtain the test solution; (b) Preparation of mixed reference solution: accurately weigh hesperidin, rutin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin reference substances, dilute to volume with methanol, filter, and take the filtrate to obtain a mixed reference solution; (c) Chromatographic conditions: Chromatographic column: octadecylsilane bonded silica gel as filler; mobile phase: acetonitrile (A)-0.1% formic acid solution (B); gradient elution program: 0-10 min, 10%A→15%A; 10-24 min, 15%A→25%A; 24-28 min, 25%A→26%A; 28-30 min, 26%A→30%A; 30-40 min, 30%A→70%A; 40-45 min, 70%A→10%A; 45-50 min, 10%A; dual wavelength method of 254 nm and 283 nm was used for detection, with 254 nm as the detection wavelength for amygdalin, hyperoside, quercetin, and ammonium glycyrrhizinate, and 283 nm as the detection wavelength for liquiritin, ferulic acid, hesperidin, and naringin; (d) Content determination: The mixed reference solution and the test solution were respectively injected into the HPLC, and the determination was carried out according to the above chromatographic conditions. The regression equation of each reference substance was established, and the contents of hesperidin, rutin, ammonium glycyrrhizinate, liquiritin, amygdalin, ferulic acid, hyperoside and quercetin in the test solution were calculated by the external standard method.

2. The method for determining the contents of multiple components in the Sanaoyulong mixture according to claim 1, characterized in that: In step (a), the mass concentration of the methanol is 100%.

3. The method for determining the contents of multiple components in the Sanaoyulong mixture according to claim 1, characterized in that: In step (a), the volume ratio of the Sanaoyulong mixture to methanol is 1:3-5, preferably 1:

4.

4. The method for determining the contents of multiple components in the Sanaoyulong mixture according to claim 1, characterized in that: In step (b), the mass concentration of the methanol is 75% to 85%.

5. The method for determining the contents of multiple components in Sanao Yulong mixture according to claim 1, characterized in that: In step (c), the chromatographic column is East Laboratory Epic C18, with specifications of 4.6 mm×250 mm, 5 μm.

6. The method for determining the contents of multiple components in the Sanaoyulong mixture according to claim 1, characterized in that: In step (c), the flow rate is 0.6-1.0 mL / min; the column temperature is 25-35 °C; and the injection volume is 10 μL.

7. The method for determining the contents of multiple components in Sanaoyulong mixture according to claim 1, characterized in that: In step (d), The regression equation of hesperidin is Y=7050.4X+26407, and the correlation coefficient R 2 is 0.9991; The regression equation of naringenin is Y=20156X+3093.5, and the correlation coefficient R 2 is 1; The regression equation of ammonium glycyrrhizinate is Y=9163.9X-22574, and the correlation coefficient R 2 is 0.9994; The regression equation of liquiritin is Y=19811X-587.2, and the correlation coefficient R 2 is 1; The regression equation of ferulic acid is Y=40351X+95.8, and the correlation coefficient R 2 is 1; The regression equation of amygdalin is Y = 365.2X-2461.7, and the correlation coefficient R 2 is 0.9997; The hyperoside regression equation is Y = 38283X + 358.54, and the correlation coefficient R 2 is 1; The quercetin regression equation is Y = 40127X + 11836, and the correlation coefficient R 2 is 0.9993.

8. Application of the method for determining the contents of multiple components in the Sanao Yulong mixture according to any one of claims 1 to 7 in the quality control of the Sanao Yulong mixture.

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