Quality detection method of gout clearing mixture for treating hyperuricemia

Abstract

The invention discloses a quality detection method of a gout clearing mixture for treating hyperuricemia, which adopts a separation analysis technology of ultra-high performance liquid chromatography to detect eight chemical components in the gout clearing mixture. Comprising the following components: 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neoastilbin, astilbin, berberine hydrochloride and emodin-8-beta-D-glucopyranoside. According to the method, eight chemical components of six active components in the gout clearing mixture are quantitatively analyzed at the same time, and the method has the advantages of being easy and convenient to operate, good in stability and easy to popularize. The invention can provide a comprehensive and reliable quality control basis for the gout clearing mixture, and has important significance for ensuring the clinical curative effect of the gout clearing mixture.

Description

Technical Field

[0001] The present invention belongs to the technical field of traditional Chinese medicine detection, and particularly relates to a method for simultaneously determining the contents of multiple components in Tongfeng Qingli Mixture based on UPLC-PDA technology. Background Art

[0002] Hyperuricemia is the "fourth highest" after diabetes, hypertension, and hyperlipidemia. With the changes in people's lifestyle and dietary structure, the prevalence of hyperuricemia in China has been increasing year by year and has become the second largest metabolic disease after diabetes. Clinically, the main manifestation of HUA patients is that the fasting blood uric acid level > 420 μmol / L. Hyperuricemia is not only closely related to the occurrence of gout, but also associated with chronic diseases such as cardiovascular diseases, chronic kidney diseases, obesity, and diabetes, seriously affecting people's health and quality of life.

[0003] Tongfeng Qingli Mixture is composed of 11 herbs including Smilax glabra, Polygonum cuspidatum, Phellodendron amurense, Atractylodes lancea, Dioscorea nipponica, etc. It is a hospital preparation of Yinchuan Traditional Chinese Medicine Hospital in Ningxia, which is modified based on Simiao Pills in Chengfang Biandu written by Zhang Bingcheng in the Qing Dynasty. In the formula, Smilax glabra is the monarch drug, strengthening the spleen and removing dampness, and promoting joint movement; Polygonum cuspidatum is the minister drug, clearing heat and detoxifying, resolving phlegm and promoting diuresis; Clematis chinensis, Dioscorea nipponica, and Pheretima play the role of dredging collaterals and relieving pain, Cyathula officinalis benefits the liver and kidney and guides the drugs downward, Coix lacryma-jobi removes obstruction and dampness, Phellodendron amurense, Atractylodes lancea, and Alisma orientalis clear heat and promote diuresis, and Paeonia lactiflora clears heat and relieves pain, all serving as assistant and guiding drugs. The whole formula has the effects of clearing and promoting damp-heat, discharging turbid substances and removing stasis, and dredging collaterals and relieving pain, and is applicable to the joint discomfort caused by "redness, swelling, heat, and pain", joint swelling, stabbing pain, and the formation of tophi during the acute attack period of gout. At present, there are many studies on the chemical components of single traditional Chinese medicines such as Smilax glabra, Phellodendron amurense, and Polygonum cuspidatum, but there is no research on the chemical components of Tongfeng Qingli Mixture. The efficacy of traditional Chinese medicine compounds is often achieved through the synergistic effect of multiple components. Summary of the Invention

[0004] Object of the Invention: To solve the deficiencies of the existing technology, the object of the present invention is to provide a method for detecting 8 chemical components in Tongfeng Qingli Mixture based on UPLC-PDA technology. By using reversed-phase high-performance liquid chromatography separation technology, the contents of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neoponatin, ponatin, berberine hydrochloride, and emodin-8-β-D-glucopyranoside in Tongfeng Qingli Mixture are determined. This method can simultaneously determine the contents of six types of chemical components in Tongfeng Qingli Mixture, which is of great significance for controlling its quality and ensuring its efficacy.

[0005] Technical Solution: To achieve the above object, the technical solution adopted by the present invention is as follows:

[0006] A quality detection method for Tongfeng Qingli Mixture for treating hyperuricemia, comprising the following steps:

[0007] (1) Preparation of reference substance solution

[0008] Weigh appropriate amounts of reference substances of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neofilipenduloside, filipenduloside, berberine hydrochloride, and emodin-8-β-D-glucopyranoside accurately, dissolve them in methanol and make up the volume to obtain a mixed reference substance stock solution, and then serially dilute it with methanol to prepare reference substance solutions with a series of different mass concentrations.

[0009] (2) Preparation of test sample solution

[0010] Precisely measure an appropriate amount of Tongfeng Qingli Mixture, place it in a volumetric flask, add an appropriate amount of methanol, sonicate it, and filter it through a 0.22 μm microporous membrane to obtain the test sample solution of Tongfeng Qingli Mixture.

[0011] (3) Determination of 8 chemical components in the sample

[0012] Use an ultra-high performance liquid chromatograph to detect the mixed reference substance solution and the test sample solution respectively, obtain chromatograms, and calculate the contents of various components in the mixture by the external standard curve method.

[0013] As an optimized scheme, for the quality inspection method of the above-mentioned Tongfeng Qingli Mixture for treating hyperuricemia, weigh appropriate amounts of reference substances of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neofilipenduloside, filipenduloside, berberine hydrochloride, and emodin-8-β-D-glucopyranoside accurately, dissolve them in methanol and make up the volume to prepare a mixed reference substance solution with mass concentrations of 378.00, 105.00, 540.00, 550.00, 99.00, 107.00, 54.50, and 52.50 μg / mL respectively.

[0014] As an optimized scheme, for the quality inspection method of the above-mentioned Tongfeng Qingli Mixture for treating hyperuricemia, the preparation of the test sample solution described in step (2) is as follows: precisely measure 1 mL of Tongfeng Qingli Mixture, place it in a 2 mL volumetric flask, add an appropriate amount of methanol, sonicate it, let it cool, and then make up the volume to the scale with methanol, shake well, centrifuge at 10000 - 13000 rpm·min -1 for 10 - 15 min, take the supernatant and filter it through a 0.22 μm microporous membrane to obtain it.

[0015] As an optimized scheme, for the quality inspection method of the above-mentioned Tongfeng Qingli Mixture for treating hyperuricemia,

[0016] The conditions for the above-mentioned ultra-high performance liquid chromatography determination are as follows:

[0017] Chromatographic column: The chromatographic column model is ACQUITY UHPLC BEH C 18Chromatographic column, specifications: 2.1 mm × 100 mm, 1.7 μm;

[0018] Mobile phase: An aqueous solution of formic acid with a volume concentration of 0.1% is used as phase A, and acetonitrile is used as phase B for gradient elution;

[0019] Flow rate: 0.1 mL / min to 0.5 mL / min;

[0020] Column temperature: 25 °C to 35 °C;

[0021] Detection wavelength: 200 - 400 nm.

[0022] As a preferred embodiment, the quality detection method of the Tongfeng Qingli mixture for treating hyperuricemia described above,

[0023] The gradient elution program of the mobile phase is as follows: 0 - 10 min, 2% - 5% B; 10 - 25 min, 5% - 8% B; 25 - 40 min, 8% - 15% B; 40 - 48 min, 15% - 18% B; 48 - 56 min, 18% - 30% B; 56 - 59 min, 30% - 95% B; 59 - 60 min, 95% - 2% B.

[0024] As a preferred embodiment, the quality detection method of the Tongfeng Qingli mixture for treating hyperuricemia described above is characterized in that the flow rate is: 0.4 mL / min, the column temperature is: 30 °C, and the injection volume is: 2 μL.

[0025] As a preferred embodiment, the quality detection method of the Tongfeng Qingli mixture for treating hyperuricemia described above, the detection wavelength in step (3) is: 3 - O - feruloylquinic acid, 5 - O - caffeoylshikimic acid 326 nm, paeoniflorin 230 nm, polydatin, neoponatin, ponatin 291 nm, berberine, emodin - 8 - β - D - glucopyranoside 260 nm.

[0026] The present invention selects 8 chemical components, namely 3 - O - feruloylquinic acid, 5 - O - caffeoylshikimic acid, paeoniflorin, polydatin, neoponatin, ponatin, berberine hydrochloride, and emodin - 8 - β - D - glucopyranoside, which are closely related to the treatment of hyperuricemia by the Tongfeng Qingli mixture, have relatively high contents, and are representative, as the determination indexes.

[0027] The 3-O-feruloylquinic acid and 5-O-caffeoylshikimic acid determined in the present invention are organic acid components, paeoniflorin is a monoterpene component, polydatin is a stilbene component, neofilipenduloside and filipenduloside are flavonoid components, berberine is an alkaloid component, and emodin-8-β-D-glucopyranoside is an anthraquinone component. Therefore, a total of 4 detection wavelengths, namely 326 nm, 230 nm, 291 nm, and 260 nm, were selected.

[0028] The present invention optimized the chromatographic conditions:

[0029] 1. Optimization of the flow rate

[0030] The present invention investigated two different flow rates of 0.3 mL / min and 0.4 mL / min. As Figure 5 and 6 shown, at a flow rate of 0.4 mL / min, the peak shape is the best, and the resolution of each component can reach basic separation.

[0031] 2. Optimization of the gradient elution program

[0032] Elution gradient 1:

[0033] Chromatographic column: ACQUITY UHPLC BEH C 18 Chromatographic column (2.1 mm × 100 mm, 1.7 μm); the mobile phase was 0.1% formic acid in water (A) - acetonitrile (B); the gradient elution program was as follows: 0 - 15 min, 2% - 5% B; 15 - 25 min, 5% - 8% B; 25 - 40 min, 8% - 15% B; 40 - 48 min, 15% - 18% B; 48 - 56 min, 18% - 30% B; 56 - 59 min, 30% - 95% B; 59 - 60 min, 95% - 2% B. Flow rate: 0.4 mL / min. Column temperature: 30 °C. Injection volume: 2 μL. The detection wavelengths for content determination were 326 nm (3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid), 230 nm (paeoniflorin), 291 nm (polydatin, neofilipenduloside, filipenduloside), and 260 nm (berberine, emodin-8-β-D-glucopyranoside). As Figure 6 shown, under this elution condition, some compounds still did not reach baseline separation, and the elution ratio was further adjusted.

[0034] Elution gradient 2:

[0035] Chromatographic column: ACQUITY UHPLC BEH C 18Chromatographic column (2.1 mm × 100 mm, 1.7 μm); mobile phase: 0.1% formic acid in water (A) - acetonitrile (B); gradient elution program: 0 - 10 min, 2% - 5% B; 10 - 25 min, 5% - 8% B; 25 - 40 min, 8% - 15% B; 40 - 48 min, 15% - 18% B; 48 - 56 min, 18% - 30% B; 56 - 59 min, 30% - 95% B; 59 - 60 min, 95% - 2% B. Flow rate: 0.4 mL / min. Column temperature: 30 °C. Injection volume: 2 μL. Detection wavelengths for content determination: 326 nm (3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid), 230 nm (paeoniflorin), 291 nm (polydatin, neoponatin, ponatin), 260 nm (berberine, emodin-8-β-D-glucopyranoside). As Figure 7 shown, under these elution conditions, the target compounds achieved baseline separation, and had good peak shapes, meeting the requirements for quantification.

[0036] The gout-relieving and dampness-clearing mixture described in the present invention is an oral liquid prepared from smilax glabra, polygonum cuspidatum, clematis chinensis, dioscorea nipponica, pheretima, cyathula officinalis, coix lacryma-jobi, phellodendron amurense, atractylodes lancea, alisma orientale, and paeonia lactiflora.

[0037] Compared with the prior art, the present invention has the following beneficial effects:

[0038] The method for detecting the contents of chemical components in the gout-relieving and dampness-clearing mixture provided by the present invention can effectively detect the contents of a total of 8 components, namely 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neoponatin, ponatin, berberine hydrochloride, and emodin-8-β-D-glucopyranoside, simultaneously. The detection steps are simple, and it has good linearity, repeatability, and stability, which can provide a comprehensive and reliable quality control basis for the gout-relieving and dampness-clearing mixture, and is of great significance for ensuring its clinical efficacy. Description of the Drawings

[0039] Figure 1 UPLC chromatograms of the mixed reference substances and the gout-relieving and dampness-clearing mixture at 230 nm;

[0040] Figure 2 UPLC chromatograms of the mixed reference substances and the gout-relieving and dampness-clearing mixture at 326 nm;

[0041] Figure 3 UPLC chromatograms of the mixed reference substances and the gout-relieving and dampness-clearing mixture at 291 nm;

[0042] Figure 4 UPLC chromatograms of the mixed reference substances and the gout-relieving and dampness-clearing mixture at 260 nm.

[0043] Figure 5Results of the investigation at a flow rate of 0.3 mL / min;

[0044] Figure 6 Results of the investigation at a flow rate of 0.4 mL / min;

[0045] Figure 7 Results of the investigation of different gradient elution programs.

[0046] The above Figures 1 to 4 In the above, A is the chromatogram of Tongfeng Qingli Mixture; B is the chromatogram of the mixed reference substances; Peak 1. 3-O-feruloylquinic acid; Peak 2. 5-O-caffeoylshikimic acid; Peak 3. Paeoniflorin; Peak 4. Polydatin; Peak 5. Neofilipenduloside; Peak 6. Filipenduloside; Peak 7. Berberine; Peak 8. Emodin-8-β-D-glucopyranoside. Detailed implementation mode

[0047] The present invention will be further illustrated below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. After reading the present invention, various equivalent modifications made by those skilled in the art to the present invention all fall within the scope defined by the appended claims of this application.

[0048] Example: A quality detection method for Tongfeng Qingli Mixture for treating hyperuricemia.

[0049] Reagents and materials used in this example

[0050] 1 Experimental reagents

[0051] Reference substances 3-O-feruloylquinic acid (batch number: CFS202301), 5-O-caffeoylshikimic acid (batch number: CFS202301), polydatin (batch number: CFS202301), neofasciclin I (batch number: CFS202302), fasciclin I (batch number: CFS202301), emodin-8-β-D-glucopyranoside (batch number: CFS202301) were purchased from Wuhan Tianzhi Company. Berberine hydrochloride (batch number: 110713-202015, purity 85.9%) was purchased from the National Institutes for Food and Drug Control. Paeoniflorin (batch number: M28GB143089) was purchased from Shanghai Yuanye Bio-Technology Co., Ltd. Except for berberine hydrochloride, the purity of all the above reference substances was ≥98%. The 7 batches of Tongfeng Qingli Mixture were all provided by Ningxia Yinchuan Hospital of Traditional Chinese Medicine (the approval number for hospital preparations was: Ningyao Zhubei Zi Z20240012000), tfql015-sjh02-05 (S1), tfql015-xhx06-08 (S2), tfql015-ml07-09 (S3), tfql015-jwz6-17 (S4), tfql015-zjt7-18 (S5), tfql015-zjt-3 (S6). Methanol, acetonitrile, and formic acid were all chromatographically pure; the experimental water was Watsons drinking water.

[0052] 2 Methods and Results

[0053] 2.1 Preparation of reference substance solutions

[0054] Appropriately weigh the reference substances of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neofasciclin I, fasciclin I, berberine hydrochloride, and emodin-8-β-D-glucopyranoside, accurately weigh them, dissolve them with methanol and make up the volume to prepare a mixed reference substance solution with mass concentrations of 378.00, 105.00, 540.00, 550.00, 99.00, 107.00, 54.50, and 52.50 μg / mL respectively, and serially dilute it to prepare 7 reference substance solutions with different mass concentrations.

[0055] 2.2 Preparation of test sample solutions

[0056] Precisely measure 1 mL of Tongfeng Qingli Mixture, place it in a 2-mL volumetric flask, add an appropriate amount of methanol, ultrasonicate (400 W, 40 kHz) for 5 min, cool it down, and make up the volume to the scale with methanol, shake well, centrifuge at 13000 rpm·min -1 for 10 min, take the supernatant and filter it through a 0.22-μm microporous membrane to obtain the test sample solution.

[0057] 2.3 Chromatographic conditions

[0058] ACQUITY UHPLC BEH C 18Chromatographic column (2.1 mm × 100 mm, 1.7 μm); mobile phase: 0.1% formic acid in water (A) - acetonitrile (B); gradient elution program: 0 - 10 min, 2% - 5% B; 10 - 25 min, 5% - 8% B; 25 - 40 min, 8% - 15% B; 40 - 48 min, 15% - 18% B; 48 - 56 min, 18% - 30% B; 56 - 59 min, 30% - 95% B; 59 - 60 min, 95% - 2% B. Flow rate: 0.4 mL / min. Column temperature: 30 °C. Injection volume: 2 μL. Detection wavelengths for content determination: 326 nm (3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid), 230 nm (paeoniflorin), 291 nm (polydatin, neoponiferin, poniferin), 260 nm (berberine, emodin-8-β-D-glucopyranoside).

[0059] 2.4 Methodological investigation

[0060] 2.4.1 Investigation of linear relationship

[0061] Take the mixed reference substance solution prepared by the method under "2.1", inject and analyze according to the chromatographic conditions under "2.3", and record the peak areas. Using the peak areas of each reference substance as the ordinate (Y) and the mass concentrations of each reference substance as the abscissa (X), draw a standard curve. The linear relationships are shown in Table 1.

[0062] Table 1 Linear relationships of 8 components in Tongfeng Qingli Mixture

[0063]

[0064]

[0065] 2.4.2 Precision test

[0066] Take the mixed reference substance solution with medium mass concentration, inject continuously 6 times according to the chromatographic conditions under "2.3", and determine the peak areas. Calculate the RSDs of the peak areas of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neoponiferin, poniferin, berberine hydrochloride, and emodin-8-β-D-glucopyranoside to be 2.20%, 1.00%, 2.92%, 0.85%, 0.88%, 0.87%, 4.74%, and 0.99% respectively. The results show that the precision of the instrument is good.

[0067] 2.4.3 Stability test

[0068] Take the same batch of test samples (S4), inject and analyze them at 0, 2, 4, 8, 12, and 24 h according to the chromatographic conditions under "2.3", and measure the peak areas. Calculate the RSDs of the peak areas of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neofilipenduloside, filipenduloside, berberine hydrochloride, and emodin-8-β-D-glucopyranoside to be 1.55%, 3.10%, 4.10%, 2.47%, 1.23%, 1.60%, 4.35%, and 3.66% respectively. The results show that the test sample solution has good stability within 24 h.

[0069] 2.4.4 Repeatability experiment

[0070] Take 6 portions of the test sample solution (S4) prepared according to the method under "2.2" from the same batch of test samples, inject and analyze them according to the chromatographic conditions under "2.3", measure the peak areas and calculate the contents of 8 components. Calculate the RSDs of the contents of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neofilipenduloside, filipenduloside, berberine hydrochloride, and emodin-8-β-D-glucopyranoside to be 0.70%, 1.24%, 4.09%, 1.04%, 0.39%, 0.36%, 2.98%, and 2.55% respectively. The results show that the method has good repeatability.

[0071] 2.4.5 Spiked recovery experiment

[0072] Take the samples of Tongfeng Qingli Mixture (S4) with known contents, add equal amounts of single reference substance solutions (using methanol as the solvent) respectively, prepare 6 portions of the test sample solution according to the method under "2.2", inject and analyze them according to the chromatographic conditions under "2.3", measure the peak areas and calculate the spiked recoveries. Calculate the average spiked recoveries of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neofilipenduloside, filipenduloside, berberine hydrochloride, and emodin-8-β-D-glucopyranoside to be 104.50%, 102.14%, 100.40%, 103.44%, 98.33%, 98.86%, 102.22%, and 101.58%. The RSDs are 0.31%, 2.28%, 3.17%, 0.78%, 1.33%, 1.73%, 2.90%, and 3.15% respectively. The results show that the method has good accuracy.

[0073] 2.5 Determination of sample content

[0074] Take 6 batches of Tongfeng Qingli Mixture liquid medicines, prepare 3 portions of the test sample solution in parallel for each batch according to the method under "2.2", inject and determine according to the chromatographic conditions under "2.3", and determine the contents of 8 components in the samples. The results are shown in Table 2 and Table 3.

[0075] Table 2 Determination results of 8 components in each batch of Tongfeng Qingli Mixture (μg / mL, n = 3)

[0076]

[0077] Table 3 Determination results of 8 components in each batch of Tongfeng Qingli Mixture (μg / mL, n = 3)

[0078]

[0079] The above are only the preferred embodiments of the present invention. It should be pointed out that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A quality detection method for Tongfeng Qingli Mixture for treating hyperuricemia, characterized in that, It includes the following steps: (1) Preparation of reference substance solution Weigh appropriate amounts of reference substances of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neoponatin, ponatin, berberine hydrochloride, and emodin-8-β-D-glucopyranoside accurately, dissolve them in methanol and make up the volume to a constant volume to prepare a mixed reference substance stock solution, and then serially dilute it with methanol to prepare reference substance solutions with a series of different mass concentrations. (2) Preparation of test sample solution Precisely measure an appropriate amount of Tongfeng Qingli Mixture, place it in a volumetric flask, add an appropriate amount of methanol, sonicate it, and filter it through a 0.22 μm microporous filter membrane to obtain the test sample solution of Tongfeng Qingli Mixture. (3) Determination of 8 chemical components in the sample Use an ultra-high performance liquid chromatograph to detect the mixed reference substance solution and the test sample solution respectively to obtain chromatograms, and calculate the contents of various components in the mixture by the external standard standard curve method.

2. The quality detection method of the Tongfeng Qingli mixture for treating hyperuricemia according to claim 1, characterized in that, Weigh appropriate amounts of reference substances of 3-O-feruloylquinic acid, 5-O-caffeoylshikimic acid, paeoniflorin, polydatin, neoponatin, ponatin, berberine hydrochloride, and emodin-8-β-D-glucopyranoside accurately, dissolve them in methanol and make up the volume to a constant volume to prepare a mixed reference substance solution with mass concentrations of 378.00, 105.00, 540.00, 550.00, 99.00, 107.00, 54.50, and 52.50 μg / mL respectively.

3. The quality detection method of the Tongfeng Qingli mixture for treating hyperuricemia according to claim 1, characterized in that, The preparation of the test solution described in step (2) is as follows: precisely measure 1 mL of Tongfeng Qingli Mixture, place it in a 2-mL volumetric flask, add an appropriate amount of methanol, sonicate, allow it to cool, then make up the volume to the mark with methanol, shake well, centrifuge at 10000 - 13000 rpm·min -1 for 10 - 15 min, take the supernatant and filter it through a 0.22-μm microporous membrane to obtain the solution.

4. The quality inspection method of the Tongfeng Qingli Mixture for treating hyperuricemia according to claim 1, characterized in that The conditions for the above ultra-high performance liquid chromatography determination are as follows: Chromatographic column: The chromatographic column model is ACQUITY UHPLC BEH C 18 chromatographic column, with a specification of 2.1 mm × 100 mm, 1.7 μm; Mobile phase: Take an aqueous solution of 0.1% formic acid by volume as phase A and acetonitrile as phase B, and perform gradient elution. Flow rate: 0.1 mL / min to 0.5 mL / min; Column temperature: 25 °C to 35 °C; Detection wavelength: 200 - 400 nm.

5. The quality inspection method of the Tongfeng Qingli Mixture for treating hyperuricemia according to claim 4, characterized in that The gradient elution program of the mobile phase is as follows: 0 - 10 min, 2% - 5% B; 10 - 25 min, 5% - 8% B; 25 - 40 min, 8% - 15% B; 40 - 48 min, 15% - 18% B; 48 - 56 min, 18% - 30% B; 56 - 59 min, 30% - 95% B; 59 - 60 min, 95% - 2% B.

6. The quality inspection method of the Tongfeng Qingli mixture for treating hyperuricemia according to claim 4, characterized in that, The flow rate is: 0.4 mL / min, the column temperature is: 30 °C, and the injection volume is: 2 μL.

7. The quality inspection method of the Tongfeng Qingli mixture for treating hyperuricemia according to claim 1, characterized in that, The detection wavelength in step (3) is: 326 nm for 3-O-feruloylquinic acid and 5-O-caffeoylshikimic acid, 230 nm for paeoniflorin, 291 nm for polydatin, neoponatin, and ponatin, and 260 nm for berberine and emodin-8-β-D-glucopyranoside.

8. The quality inspection method of the Tongfeng Qingli mixture for treating hyperuricemia according to claim 1, characterized in that, Preparation of the standard curve: Take the mixed reference substance solution described in claim 2, serially dilute it to prepare 7 reference substance solutions with different mass concentrations, inject them into the ultra-high performance liquid chromatograph, use the concentration as the abscissa and the peak area as the ordinate to plot the standard curve equation as follows:

9. The quality inspection method of the Tongfeng Qingli mixture for treating hyperuricemia according to claim 1, characterized in that, Gout-Clearing and Dampness-Resolving Mixture is an oral liquid prepared from Smilax glabra Roxb., Polygonum cuspidatum Sieb. et Zucc., Clematis chinensis Osbeck, Dioscorea nipponica Makino, Pheretima aspergillum (E. Perrier), Cyathula officinalis Kuan, Coix lacryma-jobi L., Phellodendron amurense Rupr., Atractylodes lancea (Thunb.) DC., Alisma orientale (Sam.) Juz. and Paeonia lactiflora Pall.

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