Extraction method and content determination method of scopolamine and hyoscyamine in xingqi tantanikal capsule

Abstract

The application discloses a kind of extraction methods of scopolamine and hyoscyamine in Qiangqi Tannicar Capsules and content determination method.The extraction method comprises the following steps: Qiangqi Tannicar Capsules and the filtrate obtained by mixing first solvent are mixed, dried, and the residue is obtained;The first solvent is an alcohol solution containing acid;The filtrate obtained by mixing the residue with the second solvent is placed in an activated solid-phase extraction column, and is eluted by washing and eluting;The eluent of the solid-phase extraction column is a mixture of acetonitrile and concentrated ammonia water;The volume ratio of acetonitrile and concentrated ammonia water is 75-98:2-25.The application uses solid-phase extraction technology to avoid using chloroform, reduces the risk in the content determination process, and solid-phase extraction only needs 20 minutes.The total extraction time can be 35-65 minutes.The application realizes the enrichment and separation of scopolamine and hyoscyamine, provides a reference for the establishment of quality standards for Qiangqi Tannicar Capsules, and has important significance for the quality control of the variety.

Description

Technical Field

[0001] This invention relates to a method for extracting and determining the content of hyoscyamine and scopolamine in Xingqi Tannical capsules. Background Technology

[0002] The Uyghur medicine Xingqi Tanikar Capsules are listed in the first volume of the Uyghur Medicine section of the "Drug Standards of the Ministry of Health of the People's Republic of China". They are made from four medicinal herbs: aloe vera, pepper, borax, and daphne genkwa. They have the effects of regulating qi, relieving constipation, and alleviating pain, and are clinically used for symptoms such as loss of appetite, abdominal distension, and constipation. Daphne genkwa (Hyoscyamus niger L.) is an important component of this formula, possessing antispasmodic, analgesic, antiasthmatic, and sedative effects, and is mainly used to treat stomach cramps, cough, asthma, and mania. Scopolamine and hyoscyamine are the active and toxic components of Datura stramonium. They can effectively block central M-cholinergic receptors, inhibit the central nervous system, and exert sedative and analgesic effects. However, overdose can lead to toxic reactions such as dry mouth, difficulty swallowing, headache, and tachycardia. Therefore, the content of these two components in Xingqi Tannical capsules needs to be effectively controlled. The Chinese Pharmacopoeia also stipulates that the total content of scopolamine and hyoscyamine in Datura stramonium should not be less than 0.08%.

[0003] The current standard for Xingqi Tannikar capsules only includes thin-layer chromatography (TLC) identification of aloin and piperine. Luo Ji et al. established a TLC identification method for the medicinal materials in Xingqi Tannikar capsules and a titration method for the content of borax (Luo Yuan, Fang Luyan, Wang Jianfen, et al. Identification and determination of borax in Xingqi Tannikar capsules [J]. West China Pharmaceutical Journal, 2008(02):237-238). Pan Weicen et al. established a method for determining the content of aloin and piperine in the capsules (Pan Weiqin, Zhang Qingli, Liu Xiaofang, et al. HPLC method for determining the content of aloin in Uyghur medicine Xingqi Tannikar capsules [J]. Journal of Pharmaceutical Analysis, 2010,30(05):933-935). However, no method for determining the content of hyoscyamine and scopolamine in Xingqi Tannikar capsules has been reported. Furthermore, the pepper in the prescription of Xingqi Tannikar capsules contains a large amount of alkaloids, which greatly interferes with the determination of the content of the target components.

[0004] Currently, existing technologies have reported methods for determining the content of hyoscyamine and scopolamine in other drugs. For example, the Chinese Pharmacopoeia discloses an extraction method for hyoscyamine and scopolamine from Datura stramonium. This method involves extraction with organic solvents, alkalization with ammonia, and multiple extractions with chloroform. This method causes serious environmental hazards and physiological toxicity.

[0005] Therefore, there is an urgent need to provide a safe, sensitive, and convenient method for the extraction and content determination of hyoscyamine and scopolamine from Xingqi Tannical capsules. Summary of the Invention

[0006] The technical problem this invention aims to solve is to overcome the shortcomings of existing technologies where scopolamine and hyoscyamine are mostly extracted using organic solvents containing chloroform, causing serious environmental hazards and physiological toxicity. This invention provides a method for extracting and determining the content of scopolamine and hyoscyamine in Xingqi Tannical capsules. This invention uses solid-phase extraction technology, which avoids the use of chloroform, greatly reducing the danger in the content determination process, and the solid-phase extraction only requires 20 minutes. The total extraction time of this invention is short, only 35-65 minutes. This invention uses high-performance liquid chromatography to achieve the enrichment and separation of scopolamine and hyoscyamine, providing a reference for establishing quality standards for Xingqi Tannical capsules and having significant implications for the quality control of this product.

[0007] The present invention solves the above-mentioned technical problems through the following technical solutions:

[0008] This invention provides a method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsule, which includes the following steps:

[0009] The filtrate obtained by mixing S1 gas-releasing tannicar capsules and the first solvent is dried to obtain filter residue; the first solvent is an alcohol solution containing acid.

[0010] The filtrate obtained by mixing the filter residue described in S2 with the second solvent is placed in an activated solid-phase extraction column and eluted.

[0011] During the elution process, the eluent is a mixture of acetonitrile and concentrated ammonia; the volume ratio of acetonitrile to concentrated ammonia is (75-98):(2-25).

[0012] In step S1, the term "Qi-regulating Tannical capsule" generally refers to the contents of the Qi-regulating Tannical capsule.

[0013] In step S1, the first solvent can be obtained by mixing an acid and an alcohol. The acid can be of a type conventional in the art, preferably one or more of hydrochloric acid, sulfuric acid, and nitric acid, such as hydrochloric acid. The alcohol can be a type of alcohol conventional in the art that can dissolve the acid, such as an alcohol solvent, preferably one or more of methanol, ethanol, and isopropanol, such as methanol.

[0014] In step S1, the concentration of acid in the first solvent is preferably 0.02-0.5 mol / L, for example 0.025 mol / L, 0.05 mol / L, 0.075 mol / L, 0.1 mol / L, 0.2 mol / L, 0.3 mol / L or 0.4 mol / L.

[0015] In step S1, the acid-containing alcohol solution in the first solvent is preferably a mixture of hydrochloric acid and methanol.

[0016] In step S1, the mass-to-volume ratio of the gas-relieving tannical capsule to the first solvent can be 1g:(15-25)mL, for example, 1g:20mL.

[0017] In step S1, the mixing method can be conventional in the art, such as ultrasonic treatment.

[0018] The duration of the ultrasonic treatment can be 15-45 minutes, for example, 30 minutes.

[0019] The power of the ultrasonic treatment can be 250W. The frequency of the ultrasonic treatment can be 50kHz.

[0020] In step S1, the filtrate is obtained by filtration. The filtration is generally carried out at room temperature, which can be 10-30°C.

[0021] In step S1, the drying method can be conventional in the art, such as vacuum concentration.

[0022] In step S2, the second solvent can be of a type conventional in the art, preferably water or an acid solution. The concentration of the acid in the acid solution can be 0.025-4 mol / L, preferably 0.05-3 mol / L, for example 0.075 mol / L, 0.1 mol / L, 0.2 mol / L, 0.5 mol / L, 2 mol / L, or 3 mol / L.

[0023] Wherein, when the second solvent is an acid solution, the acid solution can be obtained by mixing the acid and the solvent. The type of acid can be conventional in the art, preferably one or more of hydrochloric acid, sulfuric acid, and nitric acid, such as hydrochloric acid. The solvent can be a conventional solvent capable of dissolving the acid, preferably water or an alcohol solvent. The alcohol solvent is preferably one or more of methanol, ethanol, and isopropanol, such as methanol.

[0024] In step S2, the mass-to-volume ratio of the gas-relieving tannicar capsule to the second solvent can be 1g:(2-6)mL, for example, 1g:4mL.

[0025] In step S2, the filtrate is obtained by filtration. The filtration is generally carried out at room temperature, which can be 10-30°C.

[0026] In step S2, those skilled in the art can determine the sample loading amount of the solid phase extraction column according to the specifications of the solid phase extraction column.

[0027] In step S2, the solid-phase extraction column is preferably a hybrid strong cation exchange solid-phase extraction column or a diatomaceous earth solid-phase extraction column. The hybrid strong cation exchange solid-phase extraction column is preferably DIKMA pro Elut PXC, TruusercoSCX, or CNW poly-Sery MCX. The diatomaceous earth solid-phase extraction column is preferably Agela Cleanert Florisil.

[0028] In step S2, the packing material of the solid phase extraction column is preferably a modified styrene-divinylbenzene copolymer, a benzenesulfonic acid-based packing material based on silica matrix, or a mixture of silica, magnesium oxide and sodium sulfate.

[0029] In step S2, the solid-phase extraction column can have specifications of 150 mg / 6 mL, 500 mg / 6 mL, 500 mg / 12 mL, or 1000 mg / 6 mL. The 150 mg, 500 mg, or 1000 mg represents the mass of the packing material in the solid-phase extraction column. The 6 mL or 12 mL represents the volume of the solid-phase extraction column.

[0030] In step S2, the solid phase extraction column is preferably a CNW poly-Sery MCX hybrid strong cation exchange solid phase extraction column, and its packing material is a modified styrene-divinylbenzene copolymer with a specification of 150 mg / 6 mL.

[0031] In step S2, the method for preparing the activated solid-phase extraction column may include the following steps: activating the solid-phase extraction column sequentially with acetonitrile and water. The volume ratio of the acetonitrile to the water is preferably (1-3):1, for example, 1:1.

[0032] In step S2, during the rinsing process, the volume ratio of the rinsing solution to the amount of sample loaded onto the solid phase extraction column can be (1-3):1, for example, 1.5:1.

[0033] In step S2, the rinsing solution is preferably any one of the following:

[0034] (1) Water and methanol;

[0035] (2) Water, methanol, and acetonitrile;

[0036] (3) Water, methanol and petroleum ether; the boiling point of the petroleum ether is preferably 30-90°C, for example 30-60°C or 60-90°C;

[0037] (4) Water, methanol and ethyl acetate.

[0038] When the rinsing solution is water and methanol, the volume ratio of water to methanol is preferably (0.5-3):1, for example, 1:1.

[0039] When the rinsing solution is water, methanol and acetonitrile, the volume ratio of water, methanol and acetonitrile is preferably (0.5-3):(0.5-3):1, for example 1:1:1.

[0040] When the rinsing solution is water, methanol and petroleum ether, the volume ratio of water, methanol and petroleum ether is preferably (0.5-3):(0.5-3):1, for example 1:1:1.

[0041] When the rinsing solution is water, methanol and ethyl acetate, the volume ratio of water, methanol and ethyl acetate is preferably (0.5-3):(0.5-3):1, for example 1:1:1.

[0042] In step S2, the volume ratio of the eluent to the filtrate solid-phase extraction column can be (0.5-3):1, for example, 1:1.

[0043] In step S2, the eluent can be prepared by mixing acetonitrile and concentrated ammonia.

[0044] In step S2, the mass percentage of the concentrated ammonia solution can be 25.0%-28.0%. The mass percentage represents the percentage of NH3 in the concentrated ammonia solution relative to the total mass of the concentrated ammonia solution.

[0045] In step S2, the volume ratio of the acetonitrile to the concentrated ammonia can be (80-98):(2-20), for example 80:20, 85:15, 90:10, 95:5, 97:3, 95:2 or 98:2, preferably (90-97):(2-10).

[0046] In step S2, the elution process preferably includes a first elution and a second elution. The volume ratio of the eluent in the first elution to the eluent in the second elution can be (0.5-3):1, for example, 1:1.

[0047] The volume ratio of acetonitrile to concentrated ammonia in the eluent of the first elution can be (90-100):2, for example, 98:2.

[0048] The volume ratio of acetonitrile to concentrated ammonia in the eluent of the second elution can be (90-100):2, for example, 95:5.

[0049] In this invention, after elution is completed, a step of drying the eluent is generally performed. This drying can be carried out by vacuum concentration.

[0050] The present invention also provides a method for determining the content of hyoscyamine and scopolamine in Xingqi Tannical capsules, which includes the following steps: detecting the test solution using a high-performance liquid chromatograph, wherein the test solution is a solution of the extract obtained by the extraction method of hyoscyamine and scopolamine in Xingqi Tannical capsules as described above.

[0051] In this invention, the high-performance liquid chromatograph can be a conventional high-performance liquid chromatograph in the art, preferably a Thermo Fisher Vanquish high-performance liquid chromatograph.

[0052] In this invention, the chromatographic column used in the high-performance liquid chromatography (HPLC) detection can be a conventional chromatographic column in the art, preferably a C18 column with an acid resistance range of pH 2-8. 18 Chromatographic columns, such as Kromasil C 18 The column is preferably 250 mm long, has an inner diameter of 4.6 mm, and has a packing particle size of 5 μm.

[0053] In this invention, the column temperature in the high-performance liquid chromatography (HPLC) detection can be conventional in the art, preferably 35°C.

[0054] In this invention, the detection wavelength in the high-performance liquid chromatography (HPLC) can be conventional in the art, preferably 210 nm.

[0055] In this invention, the mobile phase used in the high-performance liquid chromatography (HPLC) detection can be a mixture of methanol, acetonitrile, and sodium acetate solution. The flow rate of the mobile phase can be 1.0 mL / min.

[0056] The volume ratio of methanol, acetonitrile and sodium acetate solution in the mobile phase can be (14-26):(28-52):460, for example 20:40:460.

[0057] The sodium acetate solution may contain 0.1% sodium acetate by mass. This mass percentage represents the percentage of sodium acetate by mass relative to the total mass of the sodium acetate solution.

[0058] Preferably, the mobile phase also includes tetrahydrofuran. The volume percentage of the tetrahydrofuran may be 0.3%. The volume percentage represents the percentage of the volume of the tetrahydrofuran relative to the volume of the sodium acetate solution.

[0059] The pH of the mobile phase can be 6.2-6.6, for example 6.4.

[0060] The pH of the mobile phase can be adjusted using a pH adjuster, which may be glacial acetic acid.

[0061] The preparation method of the mobile phase may include the following steps: (1) first mixing sodium acetate and water to obtain a sodium acetate solution, then adding tetrahydrofuran to the sodium acetate solution to obtain a mixed solution; (2) mixing methanol, acetonitrile and the mixed solution, and then ultrasonically treating the mixture. The ultrasonic treatment time may be 15 min.

[0062] In this invention, the injection volume in the high-performance liquid chromatography (HPLC) detection can be 10 μL.

[0063] In this invention, as those skilled in the art will understand, the theoretical plate number calculated based on the atropine sulfate peak is not less than 4000 during the high performance liquid chromatography detection.

[0064] In this invention, the preparation method of the test solution preferably includes the following steps: mixing the extract obtained by the extraction method of scopolamine and hyoscyamine as described above with a solvent and filtering.

[0065] The solvent may be the mobile phase. Filtration may be performed using a filter membrane. Preferably, the pore size of the filter membrane is 0.45 μm.

[0066] In this invention, the concentration of hyoscyamine in the test solution is preferably 3.6256-45.3200 μg / mL. The concentration of hyoscyamine in the test solution is preferably 9.9920-124.9000 μg / mL.

[0067] In this invention, the preparation of a reference solution is generally performed during the content determination process. The reference solution can be prepared using conventional methods in the art, preferably comprising: accurately weighing 11.33 mg of scopolamine hydrobromide reference standard and 23.37 mg of atropine sulfate reference standard, placing them separately in 50 mL volumetric flasks, dissolving and diluting to the mark with methanol, and shaking well to obtain the reference standard stock solution. Accurately measuring an appropriate amount of the reference standard stock solution into a volumetric flask, adding methanol to prepare a mixed reference standard solution containing 45.3200 μg of scopolamine hydrobromide and 124.9000 μg of atropine sulfate per 1 mL.

[0068] In this invention, the content determination process generally includes the preparation of a negative solution. The method for preparing the negative solution preferably includes the following steps: preparing a negative solution from Xingqi Tannical capsules (containing neither scopolamine nor hyoscyamine) according to the previously described extraction method for scopolamine and hyoscyamine from Xingqi Tannical capsules.

[0069] In this invention, the contents of hyoscyamine and scopolamine in the test solution can be calculated from the contents of hyoscyamine and scopolamine in the reference solution. The contents of hyoscyamine and scopolamine in the reference solution can be obtained by plotting a standard curve and calculating from the standard curve. The plotting of the standard curve can be a conventional method in the art, preferably including: performing linear regression with the concentration of the reference solution as the abscissa and the peak area as the ordinate to plot the standard curve.

[0070] The preferred linear regression equation for scopolamine hydrobromide is Y = 0.1483X - 0.0437 (R = 1.0000), where X is preferably 3.6256 - 45.3200 μg / mL. The scopolamine content in the test solution can be calculated as: scopolamine hydrobromide content in the reference solution × peak area of ​​scopolamine hydrobromide in the test solution ÷ peak area of ​​scopolamine hydrobromide in the reference solution.

[0071] The linear regression equation for atropine sulfate is preferably Y = 0.1729X - 0.3048 (R = 1.0000), where X is preferably 9.9920 - 124.9000 μg / mL. The hyoscyamine content in the test solution can be calculated as: atropine sulfate content in the reference solution × peak area of ​​hyoscyamine in the test solution ÷ peak area of ​​atropine sulfate in the reference solution.

[0072] Based on common knowledge in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0073] The reagents and raw materials used in this invention are all commercially available.

[0074] The positive and progressive effects of this invention are as follows:

[0075] 1. The extraction method of hyoscyamine and scopolamine from the capsules of Xingqi Tannical in this invention can avoid the use of chloroform, greatly reducing the danger in the content determination process. Compared with the reported methods, it is safer, more sensitive and convenient.

[0076] 2. This invention establishes a method for determining the content of scopolamine and hyoscyamine, which realizes the enrichment and separation of scopolamine and hyoscyamine, providing a reference for the establishment of quality standards for Xingqi Tannical Capsules, and is of great significance for the quality control of this product. Attached Figure Description

[0077] Figure 1 The chromatograms are of the test solution, negative solution, and reference solution prepared in Example 1.

[0078] Figure 2 The chromatograms are of the test solutions prepared in Examples 32-35. Detailed Implementation

[0079] Based on common knowledge in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0080] The present invention is further illustrated below by way of embodiments, but the invention is not limited to the scope of the embodiments described herein. Experimental methods in the following embodiments that do not specify specific conditions were performed according to conventional methods and conditions, or as selected according to the product instructions.

[0081] The batch numbers and sources of the raw materials and instruments used in the embodiments and comparative examples of this invention are shown in Table 1 below:

[0082] Table 1

[0083]

[0084]

[0085] Example 1

[0086] Extraction methods of scopolamine and hyoscyamine:

[0087] Take 5g of the contents of the Tannical capsules (batch number 220509) and place it in an Erlenmeyer flask. Add 100mL of 0.075mol / L hydrochloric acid-methanol solution, sonicate for 30min (250W, 50kHz), filter at room temperature, and concentrate the filtrate to dryness under reduced pressure. Dissolve the filter residue in 20 mL of 0.1 mol / L hydrochloric acid aqueous solution, filter, and take 10 mL of the filtrate into an activated solid-phase extraction column (activation steps are: first activate with 6 mL of acetonitrile, then with 6 mL of water; model: CNW poly-Sery MCX mixed strong cation exchange SPE column (150 mg / 6 mL); packing material: modified styrene-divinylbenzene copolymer). Add 10 mL of water, 10 mL of methanol, and 10 mL of petroleum ether (boiling point: 30-60℃) sequentially as eluent. Discard the eluent, and add 5 mL of a mixture of acetonitrile and concentrated ammonia (volume ratio: 98:2) and 5 mL of a mixture of acetonitrile and concentrated ammonia (volume ratio: 95:5) sequentially for elution. Collect the eluent and concentrate it to dryness under reduced pressure to obtain an extract containing scopolamine and hyoscyamine.

[0088] Methods for determining the content of scopolamine and hyoscyamine:

[0089] S1. Preparation of reference solutions: Accurately weigh 11.33 mg of scopolamine hydrobromide reference standard and 23.37 mg of atropine sulfate reference standard into separate 50 mL volumetric flasks, dissolve and dilute to the mark with methanol, and shake well to obtain the reference stock solution. Accurately measure an appropriate amount of the reference stock solution into a volumetric flask, add methanol to prepare a mixed reference solution containing 45.3200 μg of scopolamine hydrobromide and 124.9000 μg of atropine sulfate per mL.

[0090] S2. Dissolve the above-mentioned extract containing scopolamine and hyoscyamine in 2 mL of mobile phase, filter through a 0.45 μm filter membrane to obtain the test solution.

[0091] S3. Preparation of negative solution: Weigh the negative sample (batch number XQ22022101) that does not contain hyoscyamine in the prescription, and prepare a negative solution according to the extraction methods of scopolamine and hyoscyamine.

[0092] Inject the negative control solution, reference solution, and test solution into the chromatograph separately, and inject them under the following chromatographic conditions: Column: Kromasil C 18 Column (4.6 mm × 250 mm, 5 μm): Column temperature: 35℃; Mobile phase: methanol-acetonitrile-0.1% sodium acetate (containing 0.3% tetrahydrofuran, pH adjusted to 6.40 with glacial acetic acid) (20:40:460); Detection wavelength: 210 nm; Flow rate: 1.0 mL / min; Injection volume: 10 μL. The theoretical plate number, calculated based on the atropine sulfate peak, should be no less than 4000.

[0093] Example 2

[0094] The only difference from Example 1 is that the concentration of the hydrochloric acid-methanol solution in the extraction methods of scopolamine and hyoscyamine is 0.025 mol / L.

[0095] Example 3

[0096] The only difference from Example 1 is that the concentration of the hydrochloric acid-methanol solution in the extraction methods of scopolamine and hyoscyamine is 0.05 mol / L.

[0097] Example 4

[0098] The only difference from Example 1 is that the concentration of the hydrochloric acid-methanol solution in the extraction methods of scopolamine and hyoscyamine is 0.1 mol / L.

[0099] Example 5

[0100] The only difference from Example 1 is that the concentration of the hydrochloric acid-methanol solution in the extraction methods of scopolamine and hyoscyamine is 0.2 mol / L.

[0101] Example 6

[0102] The only difference from Example 1 is that the concentration of the hydrochloric acid-methanol solution in the extraction methods of scopolamine and hyoscyamine is 0.3 mol / L.

[0103] Example 7

[0104] The only difference from Example 1 is that the concentration of the hydrochloric acid-methanol solution in the extraction methods of scopolamine and hyoscyamine is 0.4 mol / L.

[0105] Example 8

[0106] The only difference from Example 1 is that in the extraction methods of scopolamine and hyoscyamine, 20 mL of water is added to the filter residue.

[0107] Example 9

[0108] The only difference from Example 1 is that in the extraction methods of scopolamine and hyoscyamine, the concentration of hydrochloric acid aqueous solution added to the filter residue is 0.025 mol / L.

[0109] Example 10

[0110] The only difference from Example 1 is that in the extraction methods of scopolamine and hyoscyamine, the concentration of hydrochloric acid aqueous solution added to the filter residue is 0.05 mol / L.

[0111] Example 11

[0112] The only difference from Example 1 is that in the extraction methods of scopolamine and hyoscyamine, the concentration of hydrochloric acid aqueous solution added to the filter residue is 0.075 mol / L.

[0113] Example 12

[0114] The only difference from Example 1 is that in the extraction methods of scopolamine and hyoscyamine, the concentration of hydrochloric acid aqueous solution added to the filter residue is 0.2 mol / L.

[0115] Example 13

[0116] The only difference from Example 1 is that in the extraction methods of scopolamine and hyoscyamine, the concentration of hydrochloric acid aqueous solution added to the filter residue is 0.5 mol / L.

[0117] Example 14

[0118] The only difference from Example 1 is that in the extraction methods of scopolamine and hyoscyamine, the concentration of hydrochloric acid aqueous solution added to the filter residue is 2 mol / L.

[0119] Example 15

[0120] The only difference from Example 1 is that in the extraction methods of scopolamine and hyoscyamine, the concentration of hydrochloric acid aqueous solution added to the filter residue is 3 mol / L.

[0121] Example 16

[0122] The only difference from Example 1 is that the eluent used in the extraction methods of scopolamine and hyoscyamine is 5 mL of water, 5 mL of methanol and 5 mL of acetonitrile.

[0123] Example 17

[0124] The only difference from Example 1 is that the eluent used in the extraction methods of scopolamine and hyoscyamine is 5 mL of water and 5 mL of methanol.

[0125] Example 18

[0126] The only difference from Example 1 is that the eluent in the extraction method of scopolamine and hyoscyamine is 5 mL water, 5 mL methanol and 5 mL petroleum ether (the boiling point of petroleum ether is 30-60℃).

[0127] Example 19

[0128] The only difference from Example 1 is that the eluent for the extraction of scopolamine and hyoscyamine is 5 mL of water, 5 mL of methanol and 5 mL of ethyl acetate.

[0129] Example 20

[0130] The only difference from Example 1 is that the eluent for the extraction of scopolamine and hyoscyamine is 5 mL of water, 5 mL of methanol and 5 mL of petroleum ether (the boiling point of petroleum ether is 60-90℃).

[0131] Example 21

[0132] The only difference from Example 1 is that the eluent in the extraction method of scopolamine and hyoscyamine is 10 mL acetonitrile: concentrated ammonia (volume ratio of 80:20).

[0133] Example 22

[0134] The only difference from Example 1 is that the eluent in the extraction method of scopolamine and hyoscyamine is 10 mL acetonitrile: concentrated ammonia (volume ratio of 85:15).

[0135] Example 23

[0136] The only difference from Example 1 is that the eluent in the extraction method of scopolamine and hyoscyamine is 10 mL acetonitrile: concentrated ammonia (volume ratio of 90:10).

[0137] Example 24

[0138] The only difference from Example 1 is that the eluent in the extraction method of scopolamine and hyoscyamine is 10 mL acetonitrile: concentrated ammonia (volume ratio of 95:5).

[0139] Example 25

[0140] The only difference from Example 1 is that the eluent in the extraction method of scopolamine and hyoscyamine is 10 mL acetonitrile: concentrated ammonia (volume ratio of 97:3).

[0141] Example 26

[0142] The only difference from Example 1 is that the eluent in the extraction method of scopolamine and hyoscyamine is 10 mL acetonitrile: concentrated ammonia (volume ratio 98:2).

[0143] Example 27

[0144] The only difference from Example 1 is that the volume of the hydrochloric acid-methanol solution used as the extraction solvent in the extraction methods of scopolamine and hyoscyamine is 50 mL.

[0145] Example 28

[0146] The only difference from Example 1 is that the volume of the hydrochloric acid-methanol solution used as the extraction solvent in the extraction methods of scopolamine and hyoscyamine is 150 mL.

[0147] Example 29

[0148] The only difference from Example 1 is that the volume of the hydrochloric acid-methanol solution used as the extraction solvent in the extraction methods of scopolamine and hyoscyamine is 200 mL.

[0149] Example 30

[0150] The only difference from Example 1 is that the ultrasonic treatment time in the extraction methods of scopolamine and hyoscyamine is 15 minutes.

[0151] Example 31

[0152] The only difference from Example 1 is that the ultrasonic treatment time in the extraction methods of scopolamine and hyoscyamine is 45 min.

[0153] Example 32

[0154] The only difference from Example 1 is that the solid phase extraction column used in the extraction method of scopolamine and hyoscyamine is DIKMApro Elut PXC (500mg / 6mL).

[0155] Example 33

[0156] The only difference from Example 1 is that the solid phase extraction column used in the extraction methods of scopolamine and hyoscyamine is DIKMApro Elut PXC (500mg / 12mL).

[0157] Example 34

[0158] The only difference from Example 1 is that the solid phase extraction column used in the extraction methods of scopolamine and hyoscyamine is Truuserco SCX (500mg / 6mL).

[0159] Example 35

[0160] The only difference from Example 1 is that the solid phase extraction column used in the extraction methods of scopolamine and hyoscyamine is AgelaCleanertFlorisil (specification 1000mg / 6mL).

[0161] Comparative Example 1

[0162] The only difference from Example 1 is that the "hydrochloric acid-methanol solution" is replaced with "methanol" in the extraction methods of scopolamine and hyoscyamine.

[0163] Comparative Example 2

[0164] The only difference from Example 1 is that the eluent in the extraction method of scopolamine and hyoscyamine is 10 mL acetonitrile: concentrated ammonia (volume ratio 99:1).

[0165] Example 1

[0166] 1. Specificity assessment

[0167] Accurately inject 10 μL each of the reference solution, test solution, and negative solution prepared in Example 1 into the high-performance liquid chromatograph, and determine the results according to the chromatographic conditions in Example 1. The results are as follows: Figure 1 See Table 2. The results show that the negative solution did not exhibit any interfering peaks at the chromatographic peaks of scopolamine hydrobromide and atropine sulfate, indicating good specificity.

[0168] Table 2 Results of Specificity Experiment

[0169]

[0170] 2. Examination of linear relationships

[0171] Prepare appropriate amounts of scopolamine hydrobromide and atropine sulfate reference standards according to the method for preparing the reference solution in Example 1, accurately weigh them, and add methanol to prepare a stock solution containing 226.6 mg and 449.6 mg per 1 mL, respectively.

[0172] (1) Accurately pipette 25 mL of scopolamine hydrobromide reference solution (stock solution) and 20 mL of atropine sulfate reference solution (stock solution) into a 100 mL volumetric flask, dilute to the mark with methanol, and shake well. (C) 氢溴酸东莨菪碱 =45.32 μg / mL, C 硫酸阿托品 =124.9μg / mL)

[0173] (2) Accurately pipette 20 mL of the mixed reference solution from step (1) into a 25 mL volumetric flask, dilute to the mark with methanol, and shake well to obtain the final product. (C) 氢溴酸东莨菪碱 =36.256 μg / mL, C 硫酸阿托品 =99.92μg / mL)

[0174] (3) Accurately pipette 20 mL of the mixed reference solution from step (2) into a 25 mL volumetric flask, dilute to the mark with methanol, and shake well to obtain the final product. (C) 氢溴酸东莨菪碱 =29.0048 μg / mL, C 硫酸阿托品 =79.936μg / mL)

[0175] (4) Accurately pipette 5 mL of the mixed reference solution from step (3) into a 10 mL volumetric flask, dilute to the mark with methanol, and shake well to obtain the final product. (C) 氢溴酸东莨菪碱 =14.50246 μg / mL, C 硫酸阿托品 =39.968μg / mL)

[0176] (5) Accurately pipette 5 mL of the mixed reference solution from step (4) into a 10 mL volumetric flask, dilute to the mark with methanol, and shake well to obtain the final product. (C) 氢溴酸东莨菪碱 =7.2512 μg / mL, C 硫酸阿托品 =19.984 μg / mL)

[0177] (6) Accurately pipette 5 mL of the mixed reference solution from step (5) into a 10 mL volumetric flask, dilute to the mark with methanol, and shake well to obtain the final product. (C) 氢溴酸东莨菪碱 =3.6256 μg / mL, C 硫酸阿托品 =9.992 μg / mL)

[0178] Accurately inject 10 μL of the above mixed reference solution into the liquid chromatograph, and record the peak area according to the chromatographic conditions in Example 1. Plot a standard curve with peak area (Y) as the ordinate and concentration (X) as the abscissa, and perform linear regression. The regression equations for scopolamine hydrobromide are Y = 0.1483X - 0.0437 (R = 1.0000) and atropine sulfate are Y = 0.1729X - 0.3048 (R = 1.0000). The results in Tables 3 and 4 show that scopolamine hydrobromide and atropine sulfate have good linearity in the concentration ranges of 3.6256-45.3200 μg / mL and 9.9920-124.9000 μg / mL, respectively.

[0179] Table 3. Linearity of Scopolamine Hydrobromide Reference Standard

[0180]

[0181]

[0182] Table 4. Linearity of Atropine Sulfate Reference Standard

[0183] Serial Number Concentration (μg / mL) Peak area 1 9.992 1.4692 2 19.984 3.1308 3 39.968 6.5791 4 79.936 13.476 5 99.92 17.01 6 124.9 21.296

[0184] 3. Precision test

[0185] Following the method for preparing the reference solution in Example 1, mixed reference solutions of scopolamine hydrobromide and atropine sulfate with mass concentrations of 14.5024 μg / mL and 39.9680 μg / mL were prepared. The solutions were injected six times consecutively. The peak area RSDs of scopolamine hydrobromide and atropine sulfate were 1.6% and 1.59% (n=6), respectively, indicating good instrument precision. The specific results are shown in Tables 5 and 6.

[0186] Six samples of the same mass with batch number 220509 were prepared as test solutions according to the preparation method of the test solution in Example 1. The samples were injected six times consecutively under the chromatographic conditions in Example 1. The results showed that the peak area RSDs of scopolamine and hyoscyamine were 1.46% and 0.63% (n=6), respectively, indicating that the instrument precision was good. The specific results are shown in Tables 5 and 6.

[0187] Table 5

[0188]

[0189]

[0190] Table 6

[0191]

[0192] 4. Repeatability test

[0193] Six samples of the same mass with batch number 220509 were prepared as test solutions according to the preparation method of the test solution in Example 1. The chromatographic conditions in Example 1 were used for determination. The peak areas of scopolamine and hyoscyamine in the six samples were measured, and the contents of scopolamine and hyoscyamine in the test solution were calculated (Note: Scopolamine content = Scopolamine hydrobromide content × 0.7894; Hyoscyamine content = Atropine sulfate content × 0.8551). The RSDs were 1.84% and 0.57%, respectively, indicating that the method has good repeatability. The results are shown in Tables 7 and 8.

[0194] Table 7 Repeatability of Scopolamine

[0195]

[0196] Table 8 Repeatability of Hyoscyamine

[0197]

[0198] 5. Intermediate precision test

[0199] Six samples of the same mass, batch number 220509, were prepared by two inspectors, A and B, from the same laboratory on different dates according to the preparation method of the test solution in Example 1. The test solutions were then analyzed under the chromatographic conditions described in Example 1. The results in Tables 9 and 10 show that the RSDs of scopolamine and hyoscyamine contents were 4.91% and 5.77% (n=12), respectively, indicating good intermediate precision of the method.

[0200] Table 9 Intermediate Precision of Scopolamine

[0201]

[0202] Table 10 Intermediate Precision of Hyoscyamine

[0203]

[0204]

[0205] 6. Stability test

[0206] The test solution was prepared from sample number 220509 according to the preparation method of the test solution in Example 1. It was stored at room temperature and measured at 0, 2, 4, 6, 8, 12 and 24 h according to the chromatographic conditions in Example 1. The peak area RSDs of scopolamine and hyoscyamine were calculated to be 1.49% and 1.71%, respectively. The results in Tables 11 and 12 show that the sample has good stability within 24 h.

[0207] Table 11 Stability of Scopolamine

[0208]

[0209] Table 12 Stability of Hyoscyamine

[0210]

[0211]

[0212] 7. Spiking recovery test

[0213] Six portions of Xingqi Tannical Capsules (batch number 220509) with known content were accurately weighed, each approximately 2.5 g. The above six portions of Xingqi Tannical Capsules were prepared into a test solution according to the preparation method of the test solution in Example 1. Then, an appropriate amount of scopolamine hydrobromide and atropine sulfate mixed reference solution were accurately added. The chromatographic conditions in Example 1 were used for determination. The results are shown in Tables 13 and 14.

[0214] Table 13 Scopolamine Recovery Test (n=6)

[0215]

[0216] Table 14 Hyoscyamine Recovery Test (n=6)

[0217]

[0218] In Tables 13 and 14, the sample weight represents the actual weight of the Xingqi Tannical capsules; the original amount represents the content of scopolamine and hyoscyamine in the Xingqi Tannical capsules (original amount = sample weight × 220509 average content of scopolamine and hyoscyamine in this batch); the added amount represents the weight of scopolamine and hyoscyamine in the mixed reference solution of scopolamine hydrobromide and atropine sulfate; the measured amount represents the content of scopolamine and hyoscyamine actually calculated using the method of this application (calculated as: measured amount = reference concentration × test sample peak area ÷ reference peak area); and the recovery rate is calculated as (measured amount - original amount) / added amount × 100%.

[0219] The results showed that the recoveries of scopolamine and hyoscyamine met the relevant requirements (85%-110%), indicating that the method is accurate and reliable.

[0220] Example 2

[0221] The test solutions of the aforementioned 10 batches of samples were prepared according to the preparation method of the test solution in Example 1. The peak areas of hyoscyamine and scopolamine in the test solutions were determined according to the chromatographic conditions of Example 1. The contents of hyoscyamine and scopolamine in the aforementioned 10 batches of samples were calculated. The results are shown in Table 15.

[0222] Table 15 Results of Sample Content Determination

[0223]

[0224] Example 2

[0225] The total content of scopolamine and hyoscyamine in the test solutions obtained in Examples 1-7 and Comparative Example 1 was determined. The average total content was obtained by taking the average value of four determinations. The determination results are shown in Table 16 below.

[0226] Table 16. Investigation of Extraction Solvents

[0227]

[0228]

[0229] Example 3

[0230] The total content of scopolamine and hyoscyamine in the test solutions obtained in Examples 1, 8-14 and Comparative Example 2 was determined. The average total content was obtained by taking the average value of the four determinations. The determination results are shown in Table 17 below.

[0231] Table 17 Investigation of Solvents for Filter Residue Dissolution

[0232] Filter residue dissolution solvent Average total content (μg / g) Example 8 water 27.1760 Example 9 0.025mol / L 21.3052 Example 10 0.050 mol / L 17.6782 Example 11 0.075 mol / L 19.7992 Example 1 0.1 mol / L 29.0615 Example 12 0.2 mol / L 27.6028 Example 13 0.5 mol / L 25.7208 Example 14 2mol / L 25.5828 Example 15 3mol / L 24.1590

[0233] Example 4

[0234] The total content of scopolamine and hyoscyamine in the test solutions obtained in Examples 1, 15-18 and Comparative Example 3 was determined. The average total content was obtained by taking the average value of the four determinations. The determination results are shown in Table 18 below.

[0235] Table 18 Investigation of Rinse Agents

[0236]

[0237] Example 5

[0238] The total content of scopolamine and hyoscyamine in the test solutions obtained in Examples 1, 19-24 and Comparative Example 4 was determined. The average total content was obtained by taking the average value of the four determinations. The determination results are shown in Table 19 below.

[0239] Table 19. Eluent Evaluation

[0240] Acetonitrile: Concentrated ammonia Average total content (μg / g) Example 21 80:20 23.3455 Example 22 85:15 20.1438 Example 23 90:10 24.8468 Example 24 95:5 25.3360 Example 25 97:3 24.7803 Example 26 98:2 23.6820 Comparative Example 2 99:1 8.7030 Example 1 95:2 95:5 26.4520

[0241] Example 6

[0242] The total content of scopolamine and hyoscyamine in the test solutions obtained in Examples 1, 25-27 was determined. The average total content was obtained by taking the average value of the four determinations. The determination results are shown in Table 20 below.

[0243] Table 20 Material Ratio Analysis

[0244] Extraction solvent dosage Average total content (μg / g) Example 27 50mL 25.1866 Example 1 100mL 37.1605 Example 28 150mL 37.1308 Example 29 200mL 38.0592

[0245] Example 7

[0246] The total content of scopolamine and hyoscyamine in the test solutions obtained in Examples 1, 28-29 was determined. The average total content was obtained by taking the average value of the four determinations. The determination results are shown in Table 21 below.

[0247] Table 21 Ultrasonic Time Assessment

[0248]

[0249]

[0250] Example 8

[0251] The total content of scopolamine and hyoscyamine in the test solutions obtained in Examples 32-35 was determined. The average total content was obtained by taking the average of four determinations. The determination results are shown in Table 22 below. Figure 2 As shown. Among them, Figure 2 a, b, c, and d represent the chromatograms of the test solutions in Examples 32, 33, 34, and 35, respectively. The retention times of scopolamine and hyoscyamine in the test solutions in Examples 32-35 are shown in Table 23.

[0252] Table 22

[0253] Solid phase extraction column models and specifications Average total content (μg / g) Example 32 DIKMA pro Elut PXC (500mg / 6mL) 21.5401 Example 33 DIKMA pro Elut PXC(500mg / 12mL) 6.6006 Example 34 Truserco SCX (500mg / 6mL) 19.4970 Example 35 AgelaCleanertFlorisil(1000mg / 6mL) 11.0402

[0254] Table 23

[0255] Retention time of scopolamine / min Retention time of hyoscyamine / min Example 32 19.277 39.483 Example 33 19.295 39.747 Example 34 19.3 39.55 Example 35 19.31 39.748

[0256] Depend on Figure 2 It can be seen that, compared with Example 1, when the model or specifications of the solid phase extraction column were changed in Examples 32-35, although the characteristic peaks of scopolamine and hyoscyamine could still be successfully separated, the resulting chromatograms contained more impurity peaks.

Claims

1. A method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsule, characterized in that, It includes the following steps: S1 The filtrate obtained by mixing the gas-releasing tannicar capsules and the first solvent is dried to obtain the filter residue; the first solvent is a methanol solution containing acid; S2 The solid-phase extraction column is activated sequentially with acetonitrile and water; the filtrate obtained by mixing the filter residue with the second solvent is placed in the activated solid-phase extraction column and eluted. The solid-phase extraction column is a CNW poly-Sery MCX mixed strong cation exchange SPE column, 150 mg / 6 mL. The second solvent is either water or an acid solution; During the rinsing process, the rinsing solution is any one of the following: (1) Water and methanol; (2) Water, methanol, and acetonitrile; (3) Water, methanol and petroleum ether; the boiling point of the petroleum ether is 30-90℃; (4) Water, methanol, and ethyl acetate; During the elution process, the eluent is a mixture of acetonitrile and concentrated ammonia; the volume ratio of acetonitrile to concentrated ammonia is (75-98):(2-25).

2. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 1, characterized in that, In step S1, the first solvent is obtained by mixing an acid and an alcohol; the acid is one or more of hydrochloric acid, sulfuric acid, and nitric acid. And / or, in step S2, when the second solvent is an acid solution, the concentration of the acid in the acid solution is 0.025-4 mol / L; And / or, in step S2, when the second solvent is an acid solution, the acid is one or more of hydrochloric acid, sulfuric acid, and nitric acid.

3. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 2, characterized in that, In step S2, when the second solvent is an acid solution, the concentration of the acid in the acid solution is 0.05-3 mol / L.

4. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 3, characterized in that, In step S2, when the second solvent is an acid solution, the concentration of the acid in the acid solution is 0.075 mol / L, 0.1 mol / L, 0.2 mol / L, 0.5 mol / L, 2 mol / L, or 3 mol / L.

5. The extraction method of hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in any one of claims 1 to 4, characterized in that, In step S1, the concentration of acid in the first solvent is 0.02-0.5 mol / L; And / or, in step S1, the mass-to-volume ratio of the gas-relieving Tannical capsule to the first solvent is 1 g: (15-25) mL; And / or, in step S2, the mass-to-volume ratio of the gas-releasing tannicar capsule to the second solvent is 1 g: (2-6) mL.

6. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 5, characterized in that, In step S1, the concentration of acid in the first solvent is 0.025 mol / L, 0.05 mol / L, 0.075 mol / L, 0.1 mol / L, 0.2 mol / L, 0.3 mol / L, or 0.4 mol / L; And / or, in step S1, the mass-to-volume ratio of the gas-relieving Tannical capsule to the first solvent is 1g:20mL; And / or, in step S2, the mass-to-volume ratio of the gas-releasing Tannical capsule to the second solvent is 1 g: 4 mL.

7. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 1, characterized in that, In step S2, during the activation of the solid-phase extraction column using acetonitrile and water in sequence, the volume ratio of acetonitrile to water is (1-3):

1.

8. The method for extracting hyoscyamine and scopolamine from the Qi-regulating Tannical capsules as described in claim 7, characterized in that, In step S2, during the activation of the solid-phase extraction column using acetonitrile and water in sequence, the volume ratio of acetonitrile to water is 1:

1.

9. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsule as described in claim 1, characterized in that, The boiling point of the petroleum ether is 30-60℃ or 60-90℃.

10. The method for extracting hyoscyamine and scopolamine from the Qi-regulating Tannical capsules as described in claim 1, characterized in that, When the rinsing solution is water and methanol, the volume ratio of water to methanol is (0.5-3):1; When the rinsing solution is water, methanol, and acetonitrile, the volume ratio of water, methanol, and acetonitrile is (0.5-3):(0.5-3):1; When the eluent is water, methanol and petroleum ether, the volume ratio of water, methanol and petroleum ether is (0.5-3):(0.5-3):1; When the eluent is water, methanol and ethyl acetate, the volume ratio of water, methanol and ethyl acetate is (0.5-3):(0.5-3):

1.

11. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 10, characterized in that, In step S2, when the rinsing solution is water and methanol, the volume ratio of water to methanol is 1:

1. When the rinsing solution is water, methanol and acetonitrile, the volume ratio of water, methanol and acetonitrile is 1:1:1; When the eluent is water, methanol and petroleum ether, the volume ratio of water, methanol and petroleum ether is 1:1:

1. When the eluent is water, methanol and ethyl acetate, the volume ratio of water, methanol and ethyl acetate is 1:1:

1.

12. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 1, characterized in that, In step S2, the concentrated ammonia solution has a mass percentage of 25.0%-28.0%. And / or, in step S2, the volume ratio of the acetonitrile to the concentrated ammonia is (80-98):(2-20); And / or, in step S2, the elution process includes a first elution and a second elution; the volume ratio of the eluent in the first elution to the eluent in the second elution is (0.5-3):1; Wherein, the volume ratio of acetonitrile and concentrated ammonia in the eluent of the first elution is (90-98):2; In the second elution, the volume ratio of acetonitrile to concentrated ammonia in the eluent is (90-98):

2.

13. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 1, characterized in that, In step S2, the volume ratio of the acetonitrile to the concentrated ammonia is 80:20, 85:15, 90:10, 95:5, 97:3, 95:2, or 98:

2.

14. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 12, characterized in that, The volume ratio of the eluent for the first elution to the eluent for the second elution is 1:

1. In the first elution, the volume ratio of acetonitrile to concentrated ammonia in the eluent is 98:

2. In the second elution, the volume ratio of acetonitrile to concentrated ammonia in the eluent is 95:

5.

15. The method for extracting hyoscyamine and scopolamine from the Xingqi Tannical capsules as described in claim 12, characterized in that, In step S2, the volume ratio of the acetonitrile to the concentrated ammonia is (90-97):(2-10).

16. A method for determining the content of hyoscyamine and scopolamine in Xingqi Tannical capsules, characterized in that, It includes the following steps: The test solution was analyzed by high performance liquid chromatography. The test solution was a solution of the extract obtained by the extraction method of hyoscyamine and scopolamine from Xingqi Tannical capsules according to any one of claims 1-15. In the high-performance liquid chromatography (HPLC) detection, the chromatographic column is a C18 column with an acid resistance range of pH 2-8. 18 Chromatographic column; In the high-performance liquid chromatography (HPLC) detection, the detection wavelength is 210 nm; In the high-performance liquid chromatography (HPLC) detection, the mobile phase is a mixture of methanol, acetonitrile, and sodium acetate solution; the mobile phase also includes tetrahydrofuran. In the mobile phase, the volume ratio of methanol, acetonitrile, and sodium acetate solution is (14-26):(28-52):

460.

17. The method for determining the content of hyoscyamine and scopolamine in Xingqi Tannical capsules as described in claim 16, characterized in that, The high-performance liquid chromatograph is a Thermo Fisher Vanquish high-performance liquid chromatograph. And / or, in the high-performance liquid chromatography (HPLC) detection, the chromatographic column is Kromasil C60 ... 18 The chromatographic column has the following specifications: a length of 250 mm, an inner diameter of 4.6 mm, and a packing particle size of 5 μm. And / or, in the high performance liquid chromatography detection, the column temperature is 35°C; and / or, the flow rate of the mobile phase is 1.0 mL / min.

18. The method for determining the content of hyoscyamine and scopolamine in Xingqi Tannical capsules as described in claim 17, characterized in that, The sodium acetate solution contains 0.1% sodium acetate by mass. And / or, the tetrahydrofuran has a volume percentage of 0.3%; And / or, the pH of the mobile phase is 6.2-6.

6.

19. The method for determining the content of hyoscyamine and scopolamine in Xingqi Tannical capsules as described in claim 18, characterized in that, In the mobile phase, the volume ratio of methanol, acetonitrile, and sodium acetate solution is 20:40:460; And / or, the pH of the mobile phase is 6.

4.

20. The method for determining the content of hyoscyamine and scopolamine in Xingqi Tannical capsules as described in claim 16, characterized in that, The preparation method of the test solution includes the following steps: mixing the extract obtained by the extraction method of scopolamine and hyoscyamine according to any one of claims 1-15 with a solvent and filtering; And / or, the concentration of scopolamine in the test solution is 3.6256-45.3200 μg / mL; And / or, the concentration of hyoscyamine in the test solution is 9.9920-124.9000 μg / mL; And / or, the regression equation for scopolamine hydrobromide is Y = 0.1483X - 0.0437, R = 1.0000, where X ranges from 3.6256 to 45.3200 μg / mL; where X is the concentration and Y is the peak area. And / or, the regression equation for atropine sulfate is Y = 0.1729X - 0.3048, R = 1.0000, where X ranges from 9.9920 to 124.9000 μg / mL; where X is the concentration and Y is the peak area.

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