Quality control method of rhubarb sodium bicarbonate tablets

Abstract

The invention discloses a quality control method of rhubarb sodium bicarbonate tablets. The quality control method comprises the following four parts: (1) a thin-layer chromatography TLC identification method of rhein, emodin, chrysophanol, aloe-emodin and physcion; (2) carrying out TLC (Thin Layer Chromatography) detection on rhaponticin; (3) a construction method of a high performance liquid chromatography UPLC fingerprint spectrum; and (4) a UPLC (Ultra Performance Liquid Chromatography) content determination method of aloemodin-8-0-beta-D-glucoside, rheinic acid-8-0-beta-D-glucoside, chrysophanol-8-0-beta-D-glucoside, emodin-8-0-beta-D-glucoside, emodin monoether-8-0-beta-D-glucoside, rheinic acid, emodin, chrysophanol, aloemodin and emodin monomethyl ether. The method has remarkable beneficial effects, can effectively solve the problems in the prior art, and shows excellent practical value in drug quality monitoring.

Description

Technical Field

[0001] The invention relates to the technical field of drug analysis, in particular to a quality control method for rhubarb sodium bicarbonate tablets. Background Art

[0002] The prescription of rhubarb and sodium bicarbonate tablets consists of rhubarb, sodium bicarbonate and peppermint oil, and is mainly used to improve the symptoms of excessive gastric acid, indigestion and loss of appetite. Clinical studies have shown that, in addition to the above-mentioned functions, rhubarb sodium bicarbonate tablets are also commonly used to treat other gastrointestinal diseases, kidney and cardiovascular diseases. For example, rhubarb sodium bicarbonate tablets can protect the gastric mucosa and reduce inflammatory damage (Li Jing. Clinical observation on the use of rhubarb sodium bicarbonate tablets in the treatment of critically ill children with gastrointestinal failure [J]. Journal of Pediatric Pharmacy, 2003 (3): 22-24.); can promote the recovery of gastrointestinal function after cesarean section and reduce adverse reactions (Zhou Ruihong, Shao Zhiwei, Ma Yuhui, et al. Effect of oral administration of rhubarb sodium bicarbonate tablets on gastrointestinal function recovery in cesarean section mothers [J]. Journal of Nursing, 2006 (8): 15-16.); can improve nitrogen metabolism in patients with chronic renal failure, inhibit renal compensatory hypertrophy and hypermetabolic state, and correct lipid metabolism disorders (Dai Tian, ​​Ying Fengye, Wang Lingling. Clinical observation on the treatment of chronic renal failure with rhubarb sodium bicarbonate tablets combined with alprostadil injection [J]. New Chinese Medicine, 2015, 4 7(12):73-74.); It can improve endothelial function and reduce the level of micro-inflammatory factors in the body as an auxiliary treatment for patients with uremia undergoing hemodialysis (Chen Qiaoyi, Hu Liyan, Xu Yan. Clinical study on Bailing capsule combined with rhubarb sodium bicarbonate tablets in the treatment of patients with uremia undergoing hemodialysis [J]. New Chinese Medicine, 2024, 56(6):90-94.); It can improve the nutritional status of patients with diabetic nephropathy stage III and IV and effectively delay the deterioration of renal function (Zheng Jian, Luo Wei. Study on the effect of rhubarb sodium bicarbonate tablets on the nutritional status of patients with diabetic nephropathy stage III and IV [J]. China Medical Herald, 2010, 7(28):163+168.); It can also prevent constipation in elderly patients with acute myocardial infarction and reduce the occurrence of related complications (Wang Lizi, Li Yajie, Mao Huina. Clinical observation on the effect of rhubarb sodium bicarbonate tablets in preventing constipation in elderly patients with acute myocardial infarction [J]. Nursing Research, 2003(9):533-534.). As one of the main drugs, rhubarb has the effects of purging and attacking accumulation, clearing away heat and purging fire, cooling blood and detoxifying, removing blood stasis and promoting menstruation, and removing dampness and jaundice. Pharmacological studies have shown that it has purgative, anti-tumor, anti-inflammatory, lipid regulation, kidney protection, liver protection and anti-angiogenesis effects (Xiao Xian, Li Chunyan, Xue Jintao. Research progress on the main chemical components and pharmacological effects of rhubarb [J]. Journal of Xinxiang Medical College, 2024, 41 (5): 486-496.). Therefore, when rhubarb sodium bicarbonate tablets are used to treat other gastrointestinal diseases, kidney and cardiovascular diseases, rhubarb in the prescription plays an important role.

[0003] The quality standard of Rhubarb Sodium Bicarbonate Tablets is included in the Ministry of Health Drug Standard (WS1-04(B)-89). The standard only conducts physical and chemical identification of Rhubarb Hydroxyanthraquinone and sodium bicarbonate, and content determination of sodium bicarbonate (Ministry of Health of the People's Republic of China. Drug Standard: Chemical Drugs and Preparations (Volume 1) [S]. Beijing: Medical Science and Technology Press, 1989: 8.). Up to now, there are few studies on the quality control of rhubarb sodium bicarbonate tablets. Zeng Yumei et al. used thin layer chromatography to qualitatively identify the peppermint oil in rhubarb sodium bicarbonate tablets (Zeng Yumei, Liu Zhihui, Chen Fanhua, et al. Study on the quality control of rhubarb and peppermint oil in rhubarb sodium bicarbonate tablets [J]. Chinese Drug Evaluation, 2019, 36(1): 21-25.); Hu Bo et al. used UPLC-MS / MS to determine the content of rhubarb glycosides in rhubarb sodium bicarbonate tablets (Hu Bo, Wu Xu, Wang Beibei, et al. Determination of the content of rhubarb glycosides in rhubarb sodium bicarbonate tablets and compound gentian sodium bicarbonate tablets by UPLC-MS / MS [J]. Chinese Drug Evaluation, 2020, 37(5): 362-365.); Wang Dingfeng et al. used gas chromatography to determine the content of menthol in rhubarb sodium bicarbonate tablets (Wang Dingfeng, Hu Bing. Using gas chromatography-mass spectrometry to determine the content of menthol in rhubarb sodium bicarbonate tablets). Chromatographic determination of menthol content in rhubarb sodium bicarbonate tablets [J]. Strait Pharmacy, 2022, 34(8): 64-67.); The content determination of rhubarb-related components in rhubarb sodium bicarbonate tablets mainly focuses on free anthraquinones (Wu Yong, Liu Yan, Wang Lei, et al. Quality evaluation of rhubarb sodium bicarbonate tablets based on HPLC fingerprint and one-test-multiple-evaluation method [J]. Central South Pharmacy, 2023, 21(3): 784-789; Wu Yong, Liu Yan, Chen Weidong, et al. Simultaneous determination of the contents of five components in rhubarb sodium bicarbonate tablets by HPLC [J]. Central South Pharmacy, 2019, 17(10): 1718-1720; Qin Li, Xu Xing. Determination of the contents of rhubarb-emodin and rhubarb phenol in rhubarb sodium bicarbonate tablets by HPLC [J]. Journal of Pharmaceutical Analysis, 2000, (4): 274-275.). Therefore, the following problems still exist in the quality standard or quality control research of Rhubarb Sodium Bicarbonate Tablets: (1) Only physical and chemical identification is used under the identification item, which is not specific enough; (2) The UPLC-MS / MS method is used for the detection of rhubarb glycosides, which is cumbersome; (3) The indicator components of rhubarb under the content determination item are mainly free anthraquinones of rhubarb, and rhubarb bound anthraquinones are missing, which is not comprehensive enough.

[0004] In view of this, in order to achieve effective quality control of rhubarb sodium bicarbonate tablets, the present invention establishes a qualitative and quantitative quality evaluation method, which is effective, easy to operate and stable. Summary of the invention

[0005] The invention aims to overcome the defects in the prior art that the quality control method of rhubarb sodium bicarbonate tablets cannot fully monitor the quality or the detection is time-consuming and laborious, and provides a quality control method of rhubarb sodium bicarbonate tablets with high detection efficiency, simple operation and good stability.

[0006] Aiming at the deficiencies in the prior art, the invention proposes a quality control method for rhubarb sodium bicarbonate tablets.

[0007] This is achieved specifically through the following technical solutions:

[0008] A quality control method for rhubarb sodium bicarbonate tablets, comprising four parts: (1) a thin layer chromatography (TLC) identification method for rhein, rhein, chrysophanol, aloe-rhein and physophanol methyl ether; (2) a TLC inspection method for rhein; (3) a high performance liquid chromatography (UPLC) fingerprint construction method; (4) a UPLC content determination method for aloe-rhein-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, chrysophanol-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, physophanol-8-0-β-D-glucoside, rhein, rhein, chrysophanol, aloe-rhein and physophanol methyl ether. The details are as follows:

[0009] (1) TLC Identification of Rhein, Rhein, Rhein, Aloe-Rhein and Rhein Methyl Ether

[0010] Spot the test solution A, rhein, rhein, chrysophanol, physophanol methyl ether, aloe-rhein reference solution, reference medicinal material solution and negative control solution on the same silica gel G thin layer plate, develop with developer 1, take out, dry and inspect.

[0011] in,

[0012] The test solution A is prepared according to the following steps: taking rhubarb sodium bicarbonate tablets, grinding them into powder, adding methanol aqueous solution, ultrasonicating, shaking, filtering, and obtaining the test solution A;

[0013] The rhein, rhein, chrysophanol, physophanol methyl ether, and aloe-rhein reference substance solutions are prepared according to the following steps: aloe-rhein, rhein, rhein, chrysophanol, and physophanol methyl ether are taken respectively, and dissolved in methanol aqueous solution to obtain respective reference substance solutions;

[0014] The rhubarb control medicinal material solution is prepared according to the following steps: taking rhubarb control medicinal material, treating it according to the preparation method of the test solution A, and obtaining the rhubarb control medicinal material solution;

[0015] The negative control solution is prepared according to the following steps: taking a negative sample and treating it according to the preparation method of the test solution A to obtain a negative control solution;

[0016] The developing agent 1 is a mixed solution of cyclohexane-ethyl acetate-formic acid in a volume ratio of 18:2:0.5.

[0017] Preferably, the test solution A is prepared according to the following steps: take 20 pieces of rhubarb sodium bicarbonate tablets, grind them into fine powder, take about 0.5 g, accurately weigh, put it in a conical flask, accurately add 10 mL of 70% (V / V) methanol solution, weigh the mass, ultrasonicate, cool, weigh the mass again, make up the lost mass with 70% (V / V) methanol solution, shake well, filter, and obtain the test solution A.

[0018] Preferably, the reference substance solutions of rhein, emodin, chrysophanol, physophanol methyl ether and aloe-emodin are prepared according to the following steps: appropriate amounts of aloe-emodin, rhein, emodin, chrysophanol and physophanol methyl ether are taken, accurately weighed, and dissolved in 70% (V / V) methanol solution to prepare reference substance solutions containing 0.0224, 0.0066, 0.0166, 0.0948 and 0.0848 mg per 1 mL, respectively.

[0019] Preferably, the rhubarb control medicinal material solution is prepared according to the following steps: about 0.1 g of rhubarb control medicinal material is taken and treated according to the preparation method of the test solution A in (1) to obtain the rhubarb control medicinal material solution.

[0020] Preferably, the negative control solution is prepared according to the following steps: according to the prescription ratio and preparation method of rhubarb sodium bicarbonate tablets, a negative sample lacking rhubarb medicinal materials is prepared, 0.25 g of the negative sample is taken, and treated according to the preparation method of the test solution A in (1) to obtain a negative control solution;

[0021] Preferably, the methanol aqueous solution is 70% methanol aqueous solution.

[0022] Preferably, the working conditions of the ultrasonic treatment are power 900 W, frequency 40 kHz, and time 30 min.

[0023] Preferably, the sample volume of the test solution and the negative control solution is 24 μL, the sample volume of the rhein reference solution is 21 μL, the sample volume of the rhein reference solution is 3 μL, the sample volume of the chrysophanol reference solution is 6 μL, the sample volume of the physophanol methyl ether reference solution is 18 μL, the sample volume of the aloe-emodin reference solution is 1.5 μL, and the rhubarb control medicinal material solution is 6 μL.

[0024] Preferably, the inspection conditions are inspection under ultraviolet light (365nm) and after fumigation in ammonia vapor for 3 minutes.

[0025] (2) TLC test method for rhubarb glycoside

[0026] Spot the test solution B and rhubarb glycoside reference solution on the same silica gel G thin layer plate, develop it with developer 2, take it out, dry it and inspect it.

[0027] in,

[0028] The test solution B is prepared according to the following steps: treating the test solution A according to the preparation method of (1) to obtain the test solution B;

[0029] The rhubarb glycoside reference substance solution is prepared according to the following steps: taking rhubarb glycoside, adding methanol aqueous solution to dissolve it, and obtaining the rhubarb glycoside reference substance solution;

[0030] The developing agent 2 is a mixture of chloroform-methanol-formic acid-water in a volume ratio of 100:30:2:3.

[0031] Preferably, the test solution B is prepared according to the following steps: the test solution A is treated according to the preferred preparation method of the test solution A in (1) to obtain the test solution B.

[0032] Preferably, the rhubarb glycoside reference solution is prepared according to the following steps: take the rhubarb glycoside reference, add methanol aqueous solution to dissolve it to prepare a reference solution containing 10 μg per 1 mL.

[0033] Preferably, the methanol aqueous solution is 70% methanol aqueous solution.

[0034] Preferably, the sample volume of the test solution is 6 μL, the sample volume of the rhubarb glycoside reference solution is 1.5 μL, the sample volume of the rhubarb glycoside reference solution is 3 μL, the sample volume of the chrysophanol reference solution is 6 μL, the sample volume of the physophanol methyl ether reference solution is 18 μL, the sample volume of the aloe-emodin reference solution is 1.5 μL, and the rhubarb reference medicinal material solution is 6 μL.

[0035] Preferably, the inspection condition is inspection under ultraviolet light (365nm).

[0036] (3) Construction of UPLC fingerprint

[0037] The test solution C of rhubarb sodium bicarbonate tablets and the chrysophanol reference solution were detected by ultra-high performance liquid chromatography to construct the fingerprint of rhubarb sodium bicarbonate tablets.

[0038] in,

[0039] The test solution C is prepared according to the following steps: taking rhubarb sodium bicarbonate tablets, grinding them into powder, adding 60-100% methanol aqueous solution, ultrasonicating for 30min-1h, shaking well, filtering, and obtaining the test solution C;

[0040] The chrysophanol reference substance solution is prepared according to the following steps: taking chrysophanol, adding methanol aqueous solution to dissolve it, and obtaining a reference substance solution;

[0041] The chromatographic conditions are: Waters Acquity UPLC BEH C 18 (2.1mm×100mm,1.7μm) or Agilent Edipse Plus C 18 (3.0×100mm, 1.8μm) chromatographic column, acetonitrile-0.25~1% phosphoric acid solution as mobile phase, gradient elution, detection wavelength: 222nm or 260nm, column temperature: 30~35℃, flow rate: 0.3~0.6mL / min, injection volume: 1~4μL;

[0042] Establishment of the fingerprint: inject the test solution into the liquid chromatograph for analysis and determination to obtain the chromatogram of the test solution, and import the chromatogram of the test solution into the fingerprint similarity evaluation system software for analysis to obtain the fingerprint of each batch of test samples and the final control fingerprint;

[0043] Attribution of the common peaks: attribution of the common peaks was performed by comparison with the reference substances.

[0044] Preferably, the test solution C is prepared according to the following steps: take 20 pieces of rhubarb sodium bicarbonate tablets, grind them into fine powder, take about 0.25g, accurately weigh, put it in a conical flask, accurately add 25mL of 70% (V / V) methanol solution, weigh the mass, ultrasonic (900W, 40kHz) for 30min, cool, weigh the mass again, make up the lost mass with 70% (V / V) methanol solution, shake well, filter, and obtain the test solution C.

[0045] Preferably, the chrysophanol reference solution is prepared according to the following steps: take an appropriate amount of chrysophanol, add 70% (V / V) methanol solution to dissolve it and prepare a reference solution containing 0.0229 mg per 1 mL.

[0046] Preferably, the chromatographic conditions are: Agilent Edipse Plus C 18 (3.0×100mm, 1.8μm) chromatographic column, acetonitrile (A)-0.5% phosphoric acid solution (B) as mobile phase, gradient elution: 0-3.5min, 25→65%A; 3.5-4.5min, 65→65%A; 4.5-6min, 65→90%A; 6-7min, 90→98%A; 7-7.1min, 98→25%A; 7.1-11min, 25→25%A. Detection wavelength: 222nm, column temperature: 35℃, flow rate: 0.5mL / min, injection volume: 1μL.

[0047] Preferably, the fingerprint is established by injecting 8 batches of test solution made of rhubarb sodium bicarbonate tablet powder into a liquid chromatograph for analysis and determination to obtain a chromatogram of the test solution, and importing the obtained chromatogram in .cdf format into the "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System" (2012) software for evaluation, using S1 as a reference spectrum, using the median method, and a time window width of 0.2s to process it to obtain 8 batches of test sample fingerprints and the final rhubarb sodium bicarbonate tablet control fingerprint.

[0048] Preferably, the common peak is assigned by comparing the retention time and ultraviolet absorption spectrum with those of a reference substance.

[0049] (4) UPLC content determination method for aloe-emodin-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, chrysophanol-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, physophanol-8-0-β-D-glucoside, rhein, rhein, chrysophanol, aloe-emodin and physophanol

[0050] The test solution D, mixed reference solution and negative control solution of Rhubarb Sodium Bicarbonate Tablets were analyzed by ultra-high performance liquid chromatography to determine the content.

[0051] in,

[0052] The test solution D is prepared according to the following steps: treating the test solution C according to the preparation method in (3) to obtain the test solution D;

[0053] The mixed reference substance solution is prepared according to the following steps: aloe-emodin-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, chrysophanol-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, physophanol-8-0-β-D-glucoside, physophanol-8-0-β-D-glucoside, rhein, rhein, chrysophanol, aloe-emodin and physophanol-8-0-β-D-glucoside are added with methanol aqueous solution to dissolve to obtain a mixed reference substance stock solution, and an appropriate amount of the reference substance stock solution is precisely drawn and diluted to obtain a mixed reference substance solution;

[0054] The negative control solution is prepared according to the following steps: taking a negative sample and treating it according to the preparation method of the test solution C in (3) to obtain a negative control solution;

[0055] The chromatographic conditions are: Agilent Edipse Plus C 18(3.0×100mm, 1.8μm) chromatographic column, acetonitrile-0.5% phosphoric acid solution as mobile phase, gradient elution, detection wavelength: 222nm or 260nm, column temperature: 35℃, flow rate: 0.5mL / min, injection volume: 1μL.

[0056] Preferably, the test solution D is prepared according to the following steps: treating according to the preferred preparation method of the test solution C in (3) to obtain the test solution D.

[0057] Preferably, the mixed reference solution is prepared according to the following steps: accurately weigh appropriate amounts of aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, physophanol methyl ether-8-O-β-D-glucoside, aloe-emodin, rhein, rhein, chrysophanol, physophanol methyl ether and other reference substances, place them in 5 mL volumetric flasks respectively, add Dissolve and dilute to the scale with 70% (V / V) methanol solution, shake well, and prepare reference substance stock solutions with mass concentrations of 0.460, 1.820, 0.896, 0.744, 0.362, 0.672, 0.828, 0.502, 0.572 and 0.512 mg / mL, respectively; Accurately pipette 1 mL of the reference substance stock solution, place it in a 25 mL volumetric flask, add 70% (V / V) methanol solution, make up to volume, shake well, and obtain a mixed reference substance solution.

[0058] Preferably, the negative control solution is prepared according to the following steps: according to the prescription ratio and preparation method of rhubarb sodium bicarbonate tablets, a negative sample lacking rhubarb medicinal materials is prepared, 0.125 g of the negative sample is taken, and the preparation method of the test solution C in (3) is followed to obtain a negative control solution;

[0059] Preferably, in the chromatographic conditions, the mobile phase acetonitrile (A)-0.5% phosphoric acid solution (B) gradient elution program is: 0-3.5 min, 15→37% A; 3.5-3.6 min, 37→40% A; 3.6-5 min, 40→40% A; 5-6 min, 40→60% A; 6-7 min, 60→98% A; 7-8 min, 98→98% A; 8-8.1 min, 98→15% A; 8.1-12 min, 15→15% A. Detection wavelength: 260 nm.

[0060] A quality control method for rhubarb sodium bicarbonate tablets is established by using the above methods, wherein in TLC identification and inspection, the test sample chromatogram shows the same five orange-yellow fluorescent spots at the corresponding positions of the chromatograms of aloe-emodin, rhein, rhein, chrysophanol, physophanol methyl ether reference substances and reference medicinal materials, and the spots turn red after being fumigated in ammonia vapor; the same bright blue fluorescent spots are not seen at the same positions of the chromatograms of rhubarb glycoside reference substances; UPLC fingerprints determine 20 common peaks, and attribute 10 of them to aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, and physophanol-8-O-β-D-glucoside. glucoside, chrysophanol-8-O-β-D-glucoside, physophanol methyl ether-8-O-β-D-glucoside, aloe-emodin, rhein, emodin, chrysophanol, and physophanol methyl ether; UPLC measured the contents of the 10 components in the ranges of 0.2741-2.0059, 0.5478-5.1922, 1.2092-3.9363, 1.0393-3.3302, 0.3816-1.2481, 0.1182-0.6747, 0.1680-0.9703, 0.2382-1.7893, 0.8814-4.8826, and 0.8014-5.0991 mg / g, respectively.

[0061] Compared with the prior art, the present invention has the following beneficial effects:

[0062] The invention provides a quality control method for rhubarb sodium bicarbonate tablets, which has significant beneficial effects, can effectively solve the problems existing in the prior art, and shows excellent practical value in drug quality monitoring.

[0063] First, the present invention significantly improves the accuracy and comprehensiveness of the identification and content determination of the main components in rhubarb sodium bicarbonate tablets by introducing thin layer chromatography (TLC) and high performance liquid chromatography (UPLC) technology. The traditional method is relatively single in the analysis of rhubarb components, while the present invention identifies 5 rhubarb free anthraquinone components by establishing a thin layer chromatography method, and further constructs a fingerprint of rhubarb components and a quantitative determination method of 10 rhubarb free anthraquinones and bound anthraquinone components by using the UPLC method. This method has significant advantages in the comprehensive detection of free anthraquinones and bound anthraquinone components, fills the gap in the prior art, provides a more comprehensive rhubarb component analysis method, and ensures that the quality control of the drug is more scientific and accurate.

[0064] Secondly, the present invention improves the specific identification ability of rhubarb sodium bicarbonate tablets. Although the physical and chemical identification methods used in the prior art can identify some components, their specificity is insufficient. The present invention identifies five rhubarb free anthraquinone components by TLC method, and combines the construction of UPLC fingerprint to achieve comprehensive identification of multiple key components in rhubarb sodium bicarbonate tablets, greatly improving the specificity of identification.

[0065] In addition, the quality control method of the present invention is more convenient in operation and is suitable for routine detection in large-scale production environments. Although the traditional UPLC-MS / MS method has high sensitivity, it is complicated to operate and has high cost, and is difficult to be applied on a large scale in daily quality detection. The present invention uses TLC to check rhubarb glycosides, which has a simple operation process and low detection cost, and is suitable for rapid quality monitoring in industrial production. By simplifying the operating steps, the present invention greatly improves the efficiency of quality control and reduces costs, and has significant economic benefits in particular in actual production and detection links.

[0066] Finally, the quality control method of the present invention has good stability and repeatability, and can ensure the quality consistency of different batches of Dahuang Sodium Bicarbonate Tablets. The present invention can not only provide accurate data in the detection of a single batch of products through UPLC fingerprint analysis and content determination, but also ensure the data consistency between different batches of products, effectively ensuring the quality stability of the drug.

[0067] In summary, the quality control method of the rhubarb sodium bicarbonate tablets of the present invention has the following main beneficial effects: (1) the detection method is comprehensive, covering a variety of key components such as free anthraquinone and bound anthraquinone; (2) the operation is simple, suitable for routine detection in large-scale production, and has high economic benefits; (3) the identification specificity is strong, and the key components can be effectively identified; (4) the stability and repeatability are good, ensuring the consistency of drug quality. Therefore, the present invention has important practical value in the quality monitoring of rhubarb sodium bicarbonate tablets, can effectively improve the market competitiveness of products, and has a positive role in promoting the quality standardization of the industry. BRIEF DESCRIPTION OF THE DRAWINGS

[0068] Figure 1 This is the TLC identification chromatogram of free anthraquinone of rhubarb; 1-S1; 2-S2; 3-S3; 4-S4; 5-S5; 6-S6; 7-S7; 8-S8; 9-reference medicinal material; 10-physcholine methyl ether; 11-rhein; 12-rhein; 13-aloe-emodin; 14-chrysophanol; 15-negative control;

[0069] Figure 2 Check TLC chromatogram for rhubarb glycoside; 1-S1; 2-S2; 3-S3; 4-S4; 5-S5; 6-S6; 7-S7; 8-S8; 9-rhubarb glycoside control;

[0070] Figure 3 UPLC superimposed fingerprints of 8 batches of test products and control fingerprints (R);

[0071] Figure 4 is the UPLC chromatogram of chrysophanol;

[0072] Figure 5 UPLC chromatograms of the test sample (A) and mixed reference sample (B);

[0073] Figure 6 UPLC chromatograms of the test sample (A), mixed reference sample (B) and negative control (C); 1-aloe-emodin-8-0-β-D-glucoside; 2-rheic acid-8-0-β-D-glucoside; 3-emodin-8-0-β-D-glucoside; 4-chrysophanol-8-0-β-D-glucoside; 5-physophanol methyl ether-8-0-β-D-glucoside; 6-aloe-emodin; 7-rheic acid; 8-emodin; 9-emodin; 10-physophanol methyl ether;

[0074] Figure 7 This is the TLC identification chromatogram of free anthraquinone of rhubarb [petroleum ether (30-60°C)-ethyl acetate-formic acid (15:6:1)]; 1-rhubarb methyl ether; 2-rhein; 3-rhubarb; 4-test sample; 5-reference medicinal material; 6-aloe-rhubarb; 7-rhubarb phenol;

[0075] Figure 8 This is the TLC identification chromatogram of free anthraquinone of rhubarb [cyclohexane-ethyl acetate-formic acid (12:3:0.1)]; 1-rhubarb methyl ether; 2-rhein; 3-rhubarb; 4-test sample; 5-reference medicinal material; 6-aloe-rhubarb; 7-rhubarb phenol;

[0076] Fig. 9 This is the TLC identification chromatogram of free anthraquinone of rhubarb [cyclohexane-ethyl acetate-formic acid (18:2:0.5)]; 1-rhubarb methyl ether; 2-rhein; 3-rhubarb; 4-test sample; 5-reference medicinal material; 6-aloe-rhubarb; 7-rhubarb phenol;

[0077] Fig.10 UPLC chromatograms of the test sample at 222 nm (A) and 260 nm (B). DETAILED DESCRIPTION

[0078] The technical solution of the present invention is further described below in conjunction with the accompanying drawings.

[0079] Instruments and reagents

[0080] instrument

[0081] H-Class ultra-high performance liquid chromatograph (Waters, USA); BS224S electronic balance (Sartorius, Germany); Mettler-Toledo ME203 electronic analytical balance; KQ5200E ultrasonic cleaner (Kunshan Ultrasonic Instrument Co., Ltd.); Millipore Direct-Q5UV ultrapure water system (Millipore); DZF electric constant temperature drying oven (Shanghai Jinghong Experimental Equipment Co., Ltd.); WD-9403C UV instrument (Beijing Liuyi Instrument Factory).

[0082] Reagents and drugs

[0083] Reference substances aloe-emodin-8-0-β-D-glucoside (batch number: WP23083002), rhein-8-0-β-D-glucoside (batch number: WP23082806), chrysophanol-8-0-β-D-glucoside (batch number: WP23051011), rhein-8-0-β-D-glucoside (batch number: WP2306306), physophanol-8-0-β-D-glucoside (batch number: WP23051107), rhein (batch number: WP23051206), and rhein (batch number: WP23051011). 23020110), chrysophanol (batch number: WP23052909), aloe-emodin (batch number: WP23030113) and physophanol methyl ether (batch number: WP23051107) were all purchased from Sichuan Weikeqi Biotechnology Co., Ltd. with a purity of ≥98%; rhubarb reference material (batch number: 120902-201912) was purchased from China Food and Drug Inspection Institutes; silica gel G thin layer plate was manufactured by Qingdao Ocean Chemical Co., Ltd.; cyclohexane, ethyl acetate, formic acid, chloroform, methanol, and phosphoric acid were of analytical grade, acetonitrile was of chromatographic grade, and water was ultrapure water.

[0084] Information on 8 batches of Rhubarb Sodium Bicarbonate Tablets is shown in Table 1.

[0085] Table 1 Sample information

[0086]

[0087] Example 1

[0088] A method for establishing TLC identification of free anthraquinones in rhubarb sodium bicarbonate tablets, the method comprising the following steps:

[0089] 1.1 Preparation of reference solution

[0090] Take appropriate amounts of aloe-emodin, rhein, rhein, chrysophanol, and physophanol methyl ether, weigh them accurately, add 70% (V / V) methanol aqueous solution to dissolve them and prepare reference solutions containing 0.0224, 0.0066, 0.0166, 0.0948, and 0.0848 mg per 1 mL respectively.

[0091] 1.2 Preparation of test solution

[0092] Take 20 pieces of rhubarb sodium bicarbonate tablets, grind them into fine powder, take about 0.5g, accurately weigh, put it in a conical flask, accurately add 10mL of 70% (V / V) methanol solution, weigh the mass, ultrasonic (900W, 40kHz) for 30min, cool, weigh again, make up the lost mass with 70% (V / V) methanol solution, shake well, filter to obtain the test solution.

[0093] 1.3 Preparation of control medicinal material solution

[0094] Take about 0.1 g of rhubarb reference medicinal material and prepare the reference medicinal material solution according to the method under "1.2".

[0095] 1.4 Preparation of negative control solution

[0096] According to the prescription ratio and preparation method of Rhubarb Sodium Bicarbonate Tablets, a negative sample lacking rhubarb medicinal material was prepared. Take 0.25g of the negative sample and prepare a negative control solution according to the method under "1.2".

[0097] 1.5 TLC identification of free anthraquinone

[0098] Pipette 21μL of rhein reference solution, 3μL of rhein reference solution, 6μL of chrysophanol reference solution, 18μL of physophanol methyl ether reference solution, 1.5μL of aloe-rhein reference solution, 24μL of test solution and negative control solution, and 6μL of control medicinal material solution under "1.1, 1.2, 1.3, 1.4", and spot them on the same silica gel G thin layer plate, use cyclohexane-ethyl acetate-formic acid (volume ratio 18:2:0.5) as the developing agent, develop, take out and dry. Place under ultraviolet light (365nm) for inspection, and then fumigate in ammonia vapor until the spots are clearly colored. The results showed that under ultraviolet light, in the chromatogram of the test sample, at the corresponding positions of the chromatograms of aloe-rhein, rhein, rhein, chrysophanol, physophanol methyl ether reference and control medicinal materials, the same five orange-yellow fluorescent spots appeared. Figure 1 In A, after being fumigated with ammonia vapor, the spots turned red. Figure 1 Middle B.

[0099] Example 2

[0100] A method for establishing a TLC test for rhubarb sodium bicarbonate tablets, the method comprising the following steps:

[0101] 2.1 Preparation of reference solution

[0102] Take an appropriate amount of rhubarb glycoside reference substance, add 70% (V / V) methanol solution to dissolve it to prepare a reference substance solution containing 10 μg per 1 mL.

[0103] 2.2 TLC examination of rhubarb glycosides

[0104] Pipette 1.5 μL of the rhubarb glycoside reference solution and 6 μL of the test solution under "1.2", spot them on a silica gel G thin layer plate, develop with chloroform-methanol-formic acid-water (volume ratio 100:30:2:3), and inspect under ultraviolet light (365 nm). Figure 2 The results showed that in the chromatogram of the test sample, no identical bright blue fluorescent spot was observed at the same position as in the chromatogram of the rhubarb glycoside reference sample.

[0105] Example 3

[0106] A method for constructing a UPLC fingerprint of rhubarb sodium bicarbonate tablets, the construction method comprising the following steps:

[0107] 3.1 Chromatographic conditions

[0108] Agilent Edipse Plus C 18 (3.0×100mm,1.8μm) chromatographic column, acetonitrile (A)-0.5% phosphoric acid solution (B) as mobile phase, gradient elution: 0~3.5min, 25→65%A; 3.5~4.5min, 65→65%A; 4.5~6min, 65→90%A; 6~7min, 90→98%A; 7-7.1min, 98→25%A; 7.1-11min, 25→25%A; detection wavelength: 222nm, column temperature: 35℃, flow rate: 0.5mL / min, injection volume: 1μL.

[0109] 3.2 Preparation of reference solution

[0110] Take an appropriate amount of chrysophanol, add 70% (V / V) methanol solution to dissolve it and prepare a reference solution containing 0.0229 mg per 1 mL.

[0111] 3.3 Preparation of test solution

[0112] Take 20 pieces of rhubarb sodium bicarbonate tablets, grind them into fine powder, take about 0.25g, accurately weigh, put it in a conical flask, accurately add 25mL of 70% (V / V) methanol solution, weigh the mass, ultrasonic (900W, 40kHz) for 30min, cool, weigh again, make up the lost mass with 70% (V / V) methanol solution, shake well, filter to obtain the test solution.

[0113] 3.4 Methodological Review

[0114] 3.4.1 Precision test

[0115] Take the test solution (S8) under "3.3" and inject it 6 times continuously according to the chromatographic conditions under "3.1". Take the retention time and peak area of ​​chrysophanol as reference and record the relative retention time and relative peak area of ​​each common peak. The RSD of the relative retention time of each common peak is less than 0.10% (n=6), and the RSD of the relative peak area is less than 0.42% (n=6), indicating that the precision of this method is good.

[0116] 3.4.2 Stability test

[0117] Take the test solution (S8) under item "3.3" and place it at room temperature for 0, 2, 4, 8, 12, and 24 hours. Then, inject and measure according to the chromatographic conditions under item "3.1". Take the retention time and peak area of ​​the chrysophanol peak as reference, and record the relative retention time and relative peak area of ​​each common peak. The RSD of the relative retention time of each common peak is calculated to be less than 0.76% (n=6), and the RSD of the relative peak area is less than 2.83% (n=6), indicating that each component to be tested has good stability within 24 hours.

[0118] 3.4.3 Repeatability test

[0119] Take 0.25g of sample (S8), prepare the test solution according to the method under "3.3", a total of 6 portions, and then inject and measure according to the chromatographic conditions under "3.1", take the retention time and peak area of ​​chrysophanol peak as reference, and record the relative retention time and relative peak area of ​​each common peak. The RSD of the relative retention time of each common peak is calculated to be less than 0.53% (n=6), and the RSD of the relative peak area / sample weight is less than 2.33% (n=6), indicating that this method has good repeatability.

[0120] 3.5 Ultra-high performance liquid chromatography (UPLC) fingerprint generation

[0121] Take 8 batches of rhubarb sodium bicarbonate tablets, prepare the test solution according to the method of "3.3", inject according to the chromatographic conditions under "3.1", import the obtained chromatogram in .cdf format into the "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System" (2012) software for evaluation, use S1 as the reference spectrum, use the median method, and the time window width is 0.2s to process it, match the full spectrum peaks, and confirm the common peaks. A total of 20 common peaks were determined, see Figure 3 .

[0122] 3.6 Similarity evaluation and analysis

[0123] The "Chinese Herbal Medicine Chromatographic Fingerprint Similarity Evaluation System" (2012 Edition) was used to evaluate the overall similarity with the UPLC reference fingerprint of the test sample as the reference. The results showed that the similarity of the 8 batches of test samples was greater than 0.535, indicating that the quality of Dahuang Sodium Bicarbonate Tablets from different manufacturers and batches was quite different, as shown in Table 2.

[0124] Table 2 Similarity evaluation results of 8 batches of test products

[0125]

[0126] 3.7 Relative retention time and relative peak area of ​​common peaks

[0127] By comparing with the reference solution, it was confirmed that peak 19 was chrysophanol. Figure 4 Because this peak has good separation, large peak area and stability, it is used as a reference peak to calculate the relative retention time and relative peak area of ​​other common peaks, see Tables 3 and 4.

[0128] Table 3 Relative retention time of common peaks in UPLC spectra of 8 batches of test samples

[0129]

[0130]

[0131] Table 4 Relative peak areas of common peaks in UPLC spectra of 8 batches of test samples

[0132]

[0133] 3.8 Attribution of common peaks

[0134] By comparing with the reference substance, 10 common peaks were finally assigned, peak 3 was aloe-emodin-8-0-β-D-glucoside; peak 4 was rhein-8-0-β-D-glucoside; peak 9 was rhein-8-0-β-D-glucoside; peak 10 was chrysophanol-8-0-β-D-glucoside; peak 12 was physophanol methyl ether-8-0-β-D-glucoside; peak 15 was aloe-emodin; peak 16 was rhein; peak 18 was rhein; peak 19 was chrysophanol; peak 20 was physophanol methyl ether, see Figure 5 .

[0135] Example 4

[0136] A method for establishing UPLC content determination of 10 components in rhubarb sodium bicarbonate tablets, the method comprising the following steps:

[0137] 4.1 Preparation of mixed reference solution

[0138] Accurately weigh appropriate amounts of reference substances such as aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, physophanol methyl ether-8-O-β-D-glucoside, aloe-emodin, rhein, rhein, chrysophanol, and physophanol methyl ether, and place them in 5 mL volumetric flasks, and add 70% (V / V) methanol solution. Dissolve and dilute to the scale, shake well, and prepare reference substance stock solutions with mass concentrations of 0.460, 1.820, 0.896, 0.744, 0.362, 0.672, 0.828, 0.502, 0.572 and 0.512 mg / mL, respectively; accurately pipette 1 mL of each reference substance stock solution, place it in a 25 mL volumetric flask, add 70% (V / V) methanol solution, make up to volume, shake well, and obtain a mixed reference substance solution.

[0139] 4.2 Preparation of test solution

[0140] Take 20 pieces of rhubarb sodium bicarbonate tablets, grind them into fine powder, take about 0.25g, accurately weigh, accurately add 25mL of 70% (V / V) methanol solution, weigh, ultrasonic (900W, 40kHz) for 30min, cool, weigh again, make up the lost mass with 70% (V / V) methanol solution, shake well, filter, and take the filtrate to obtain.

[0141] 4.3 Preparation of negative control solution

[0142] According to the prescription ratio and preparation method of Rhubarb Sodium Bicarbonate Tablets, a negative sample lacking rhubarb medicinal material was prepared. Take 0.125g of the negative sample and prepare a negative control solution according to the method under "4.2".

[0143] 4.4 Chromatographic conditions

[0144] Column: Agilent Edipse Plus C 18 (3.0×100mm,1.8μm), mobile phase: acetonitrile (A)-0.5% phosphoric acid solution (B), gradient elution program: 0~3.5min, 15→37%A; 3.5~3.6min, 37→40%A; 3.6~5min, 40→40%A; 5~6min, 40→60%A; 6-7min, 60→98%A; 7-8min, 98→98%A; 8-8.1min, 98→15%A; 8.1-12min, 15→15%A. Detection wavelength: 260nm, column temperature: 30℃, flow rate: 0.5mL / min, injection volume: 1μL.

[0145] 4.5 Specificity test

[0146] Take the solutions under "4.1, 4.2, 4.3" respectively, and analyze them according to the chromatographic conditions under "4.4". Figure 6 The results showed that the separation of each component was good and all were greater than 1.5, the number of theoretical plates was greater than 50,000, the retention time of each component and the corresponding peak of the reference substance was consistent, and no interference peaks appeared at the peak time of the 10 components in the negative sample, indicating good specificity.

[0147] 4.6 Methodological Review

[0148] 4.6.1 Linear relationship investigation

[0149] Take 0.3, 0.5, 1.0, 2.0, 4.0 μL of the mixed control solution in item "4.1" respectively, analyze according to the chromatographic conditions in item "4.4", and record the peak areas of the 10 components. With the mass concentration of the solution as the abscissa (X) and the peak area as the ordinate (Y), draw a standard curve and perform linear regression to calculate the linear range and correlation coefficient. The results show that the linear relationship of each component is good within the linear range, see Table 5.

[0150] Table 5 Linear regression equation, r value, linear range, detection limit and quantification limit

[0151]

[0152] 4.6.2 Limit of detection and limit of quantification

[0153] The mixed reference solution was diluted sequentially, and the injection mass concentration at a signal-to-noise ratio of 10:1 and 3:1 was used as the limit of quantification (LOQ) and limit of detection (LOD), respectively. The results are shown in Table 5.

[0154] 4.6.3 Precision test

[0155] Take the same sample solution (S1) and inject it 6 times continuously according to the chromatographic conditions under "4.4". The RSDs of the peak areas of aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, physophanol methyl ether-8-O-β-D-glucoside, aloe-emodin, rhein, rhein, chrysophanol, and physophanol methyl ether are 0.18%, 0.19%, 0.16%, 0.11%, 0.15%, 0.18%, 0.28%, 0.29%, 0.08%, and 0.42% (n=6), respectively. The results show that the precision of the instrument is good.

[0156] 4.6.4 Stability test

[0157] The same batch of test solution (S1) was sampled and analyzed at 0, 2, 4, 8, 12 and 24 h according to the chromatographic conditions under item "4.4". The RSDs of the peak areas of aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, physophanol methyl ether-8-O-β-D-glucoside, aloe-emodin, rhein, rhein, chrysophanol, and physophanol methyl ether were calculated to be 1.50%, 2.15%, 2.83%, 1.08%, 2.31%, 1.61%, 1.13%, 0.77%, 0.98% and 1.06% (n=6), respectively. The results showed that the test solution had good stability within 24 h.

[0158] 4.6.5 Repeatability test

[0159] Take 6 portions of the same batch of test sample (S1), prepare the test sample solution according to the method under "4.2", and analyze the sample according to the chromatographic conditions under "4.4". The RSDs of the peak areas of aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, physophanol methyl ether-8-O-β-D-glucoside, aloe-emodin, rhein, rhein, chrysophanol, and physophanol methyl ether are calculated to be 1.82%, 0.54%, 1.05%, 1.53%, 1.16%, 1.74%, 1.90%, 1.26%, 1.75%, and 2.33% (n=6), respectively. The results show that the method has good repeatability.

[0160] 4.6.6 Sample recovery test

[0161] Accurately weigh 6 portions of the same batch of Rhubarb Sodium Bicarbonate Tablets test sample (S1) with determined index component content, each portion is about 0.125 g, add the reference stock solution of each index component according to 100% of the index component content in the test sample, prepare the test sample solution according to the method under "4.2", inject and analyze according to the chromatographic conditions under "4.4", calculate the sample recovery rate, and the results are shown in Table 6.

[0162] Table 6 Test results of recovery of 10 components (n=6)

[0163]

[0164]

[0165]

[0166] 4.7 Assay

[0167] Prepare 8 batches of test solutions of Rhubarb Sodium Bicarbonate Tablets according to the method in “4.2”, and perform sample injection and determination under the chromatographic conditions in “4.4”. Calculate the contents of the 10 components respectively. The results are shown in Table 7.

[0168] Table 7 Content determination results of 8 batches of Dahuang sodium bicarbonate tablets (n=2, mg / g)

[0169]

[0170] From Table 7, we can see that the contents of aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, physophanol methyl ether-8-O-β-D-glucoside, aloe-emodin, rhein, rhein, chrysophanol, physophanol methyl ether; the contents of the 10 components were measured in the range of 0.274 1~2.0059, 0.5478~5.1922, 1.2092~3.9363, 1.0393~3.3302, 0.3816~1.2481, 0.11 82~0.6747, 0.1680~0.9703, 0.2382~1.7893, 0.8814~4.8826, 0.8014~5.0991mg / g.

[0171] Optimization of experimental conditions

[0172] Investigation of TLC conditions

[0173] Referring to the method under the quality standard identification of rhubarb slices (National Pharmacopoeia Committee. Pharmacopoeia of the People's Republic of China: Volume 1 [S]. Beijing: China Medical Science and Technology Press, 2020: 24-25.), the upper layer solution of petroleum ether (30-60°C)-ethyl acetate-formic acid (volume ratio 15:6:1) was used as the developing agent for the experiment. The results showed that in the chromatograms of the test and reference medicinal materials, the spots at the corresponding positions of the chromatograms of aloe-emodin, rhein, and rhein reference substances were well separated, but there was only one spot at the corresponding positions of the chromatograms of rhubarb methyl ether and rhubarb phenol reference substances, indicating that rhubarb methyl ether and rhubarb phenol were not separated, see Figure 7 .

[0174] Referring to the method under the identification item of the quality standard of Sanhuang Tablets (National Pharmacopoeia Committee. Pharmacopoeia of the People's Republic of China: Volume 1 [S]. Beijing: China Medical Science and Technology Press, 2020: 517-518.), cyclohexane-ethyl acetate-formic acid (volume ratio 12:3:0.1) was used as the developing solvent for the experiment. The results showed that the spots in the chromatograms of the test sample and the reference medicinal materials were well separated at the corresponding positions of the chromatograms of the other reference substances, but the separation effect of the spots in the chromatograms of the reference substances of physophanol methyl ether and chrysophanol was not ideal, see Figure 8 .

[0175] On this basis, the ratio of the developing solvent was optimized, and cyclohexane-ethyl acetate-formic acid (volume ratio 18:2:0.5) was used as the developing system. The results showed that in the chromatograms of the test and reference medicinal materials, there were spots of the same color at the corresponding positions of the chromatograms of the five rhubarb free anthraquinone reference substances, and they were well separated. Fig. 9 Therefore, cyclohexane-ethyl acetate-formic acid (volume ratio 18:2:0.5) was finally determined as the developing solvent.

[0176] Investigation of extraction conditions

[0177] The same batch of samples were extracted with 60%, 70%, 80%, 90% (V / V) methanol-water solutions and pure methanol as solvents, respectively. The samples were injected under the same chromatographic conditions and the peak areas of the 10 components were compared. It was found that the peak areas of the 7 components extracted with 70% (V / V) methanol-water solutions reached the maximum. Therefore, 70% (V / V) methanol-water solutions were determined as the extraction solvent. See Table 8.

[0178] Table 8 Comparison of extraction effects of different extraction solvents

[0179]

[0180]

[0181]

[0182] Ultrasonic extraction for 30 min and reflux extraction for 1 h were also investigated, and the results showed that the extraction rates of the 10 anthraquinone components obtained by the two methods were not much different, as shown in Table 9. Ultrasonic extraction for 30 min was short and easy to operate, so it was used as the final extraction method.

[0183] Table 9 Comparison of extraction effects of different extraction methods and time

[0184]

[0185]

[0186] Determination of Detection Wavelength for Liquid Chromatography

[0187] Combining the maximum absorption wavelengths of aloe-emodin-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, rhein-8-O-β-D-glucoside, chrysophanol-8-O-β-D-glucoside, physophanol-8-O-β-D-glucoside, aloe-emodin, rhein, rhein, chrysophanol, and physophanol, it was found that there were maximum absorptions at 222nm and 260nm. Because more chromatographic peaks were detected at a wavelength of 222nm, the detection wavelength of the fingerprint was determined to be 222nm; however, the baseline fluctuation was greater at this wavelength, so the detection wavelength for content determination was determined to be 260nm, see Fig.10 .

[0188] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, rather than to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions of the technical solution of the present invention by ordinary technicians in this field do not deviate from the essence and scope of the technical solution of the present invention.

Claims

1. A quality control method for rhubarb sodium bicarbonate tablets, characterized in that: The method consists of the following four parts: (1) thin layer chromatography (TLC) identification method of rhein, rhein, chrysophanol, aloe-rhein and physophanol methyl ether; (2) TLC inspection method of rhein; (3) UPLC fingerprint construction method; (4) UPLC content determination method of aloe-rhein-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, chrysophanol-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, physophanol-8-0-β-D-glucoside, rhein, rhein, chrysophanol, aloe-rhein and physophanol methyl ether.

2. The method according to claim 1, characterized in that (1) The TLC identification method of rhein, rhein, chrysophanol, aloe-rhein and physophanol methyl ether comprises the following steps: spotting the test solution A, rhein, rhein, chrysophanol, physophanol methyl ether, aloe-rhein reference solution, rhubarb reference medicinal material solution and negative control solution on the same silica gel G thin layer plate, developing with developing agent 1, taking out, drying and inspecting.

3. The method according to claim 2, characterized in that The test solution A is prepared according to the following steps: taking rhubarb sodium bicarbonate tablets, grinding them into powder, adding methanol aqueous solution, ultrasonicating, shaking, filtering, and obtaining the test solution A; The rhein, rhein, chrysophanol, physophanol methyl ether, and aloe-rhein reference substance solutions are prepared according to the following steps: aloe-rhein, rhein, rhein, chrysophanol, and physophanol methyl ether are taken respectively, and dissolved in methanol aqueous solution to obtain respective reference substance solutions; The rhubarb control medicinal material solution is prepared according to the following steps: taking rhubarb control medicinal material, treating it according to the preparation method of the test solution A, and obtaining the rhubarb control medicinal material solution; The negative control solution is prepared according to the following steps: a negative sample lacking rhubarb is prepared according to the prescription ratio and preparation method of rhubarb sodium bicarbonate tablets, and the negative sample is taken and treated according to the preparation method of the test solution A to obtain a negative control solution; The developing agent 1 is a mixture of cyclohexane-ethyl acetate-formic acid in a volume ratio of 18:2:0.5; The methanol aqueous solution is a methanol aqueous solution with a volume percentage concentration of 70%.

4. The method according to claim 1, characterized in that: (2) The TLC test method for rhubarb glycoside comprises the following steps: spotting the test solution B and the rhubarb glycoside reference solution on the same silica gel G thin layer plate, developing with developing agent 2, taking out, drying, and inspecting.

5. The method according to claim 4, characterized in that The test solution B is prepared according to the following steps: treating the test solution A according to the preparation method of (1) to obtain the test solution B; The rhubarb glycoside reference substance solution is prepared according to the following steps: taking rhubarb glycoside, adding methanol aqueous solution to dissolve it, and obtaining the rhubarb glycoside reference substance solution; The developing agent 2 is a mixture of chloroform-methanol-formic acid-water in a volume ratio of 100:30:2:3; The methanol aqueous solution is a methanol aqueous solution with a volume percentage concentration of 70%.

6. The method according to claim 1, characterized in that (3) The construction of the UPLC fingerprint spectrum comprises the following steps: the test solution C of the rhubarb sodium bicarbonate tablets and the chrysophanol reference solution are respectively detected by ultra-high performance liquid chromatography to construct the fingerprint spectrum of the rhubarb sodium bicarbonate tablets.

7. The method according to claim 6, characterized in that The test solution C is prepared according to the following steps: take rhubarb sodium bicarbonate tablets, grind them into powder, add 70% methanol aqueous solution by volume, ultrasonicate for 30 min-1 h, shake well, filter, and obtain the test solution C; The chrysophanol reference substance solution is prepared according to the following steps: taking chrysophanol, adding methanol aqueous solution to dissolve it, and obtaining a reference substance solution; The chromatographic conditions are: Waters Acquity UPLC BEH C 18 (2.1 mm×100 mm, 1.7 μm) or Agilent Edipse Plus C 18 (3.0×100 mm, 1.8 μm) chromatographic column, acetonitrile-0.25~1% phosphoric acid solution as mobile phase, gradient elution, detection wavelength: 222 nm or 260 nm, column temperature: 30~35 ℃, flow rate: 0.3~0.6 mL / min, injection volume: 1~4 μL; Establishment of the fingerprint: inject the test solution into the liquid chromatograph for analysis and determination to obtain the chromatogram of the test solution, and import the chromatogram of the test solution into the fingerprint similarity evaluation system software for analysis to obtain the fingerprint of each batch of test samples and the final control fingerprint; Attribution of common peaks: Attribution of common peaks by comparison with reference substances; The methanol aqueous solution is a methanol aqueous solution with a volume percentage concentration of 70%.

8. The method according to claim 1, characterized in that: (4) The UPLC content determination method of aloe-emodin-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, chrysophanol-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, physophanol-8-0-β-D-glucoside, rhein, rhein, chrysophanol, aloe-emodin and physophanol comprises the following steps: the rhein sodium bicarbonate tablets test solution D, the mixed reference solution and the negative control solution are respectively analyzed by ultra-high performance liquid chromatography to determine the content.

9. The method according to claim 8, characterized in that The test solution D is prepared according to the following steps: treating the test solution C according to the preparation method in (3) to obtain the test solution D; The mixed reference substance solution is prepared according to the following steps: aloe-emodin-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, chrysophanol-8-0-β-D-glucoside, rhein-8-0-β-D-glucoside, physophanol-8-0-β-D-glucoside, physophanol-8-0-β-D-glucoside, rhein, rhein, chrysophanol, aloe-emodin and physophanol-8-0-β-D-glucoside are added with methanol aqueous solution to dissolve to obtain a mixed reference substance stock solution, and an appropriate amount of the reference substance stock solution is precisely drawn and diluted to obtain a mixed reference substance solution; The negative control solution is prepared according to the following steps: a negative sample lacking rhubarb is prepared according to the prescription ratio and preparation method of rhubarb sodium bicarbonate tablets, and the negative sample is taken and treated according to the preparation method of the test solution C in (3) to obtain a negative control solution; The chromatographic conditions are: Agilent Edipse Plus C 18 (3.0×100 mm, 1.8 μm) column, acetonitrile-0.5% phosphoric acid solution as mobile phase, gradient elution, detection wavelength: 222 nm or 260 nm, column temperature: 35 ℃, flow rate: 0.5 mL / min, injection volume: 1 μL; The methanol aqueous solution is a methanol aqueous solution with a volume percentage concentration of 70%.

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