Construction of quality control methods of radix asparagi, radix asparagi formula granules and other radix asparagi preparations

Abstract

The invention belongs to the technical field of traditional Chinese medicine detection, and particularly relates to construction of a quality control method for radix asparagi, radix asparagi formula granules and other radix asparagi preparations. Comprising the following steps: preparing a test solution; preparing a reference solution; the ultra-high performance liquid chromatography is adopted for detection, and the conditions are as follows: octadecylsilane chemically bonded silica is used as a chromatographic column of a filler; acetonitrile is used as a mobile phase A, a 0.1% phosphoric acid solution is used as a mobile phase B, and gradient elution is carried out: 0-4 min, 1%-6% A and 99%-94% B are carried out; 4-7 min, 6%-11%, 94%-89% of B; 7-12 min, 11%-15% and 89%-85% of B are carried out; and constructing a characteristic chromatogram and a control chromatogram of the radix asparagi or the preparation thereof, and comparing the characteristic chromatogram and the control chromatogram. According to the quality control method, eight common peaks of the radix asparagi or the preparation of the radix asparagi can be efficiently determined within 12 min, three characteristic peaks are identified, and rapid, environment-friendly and comprehensive quality control of radix asparagi medicinal materials, decoction and formula granules is achieved.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine testing technology, specifically involving the construction of quality control methods for Asparagus cochinchinensis, Asparagus cochinchinensis formula granules, and other Asparagus cochinchinensis preparations. Background Technology

[0002] Asparagus is the tuberous root of Asparagus cochinchinensis (Lour.) Merr., a plant in the Liliaceae family. It has the effects of nourishing yin and promoting body fluid, moistening the lungs and clearing the heart. It is mainly used to treat dry cough due to lung dryness, dry throat and thirst, internal heat and thirst, etc. It has a unique advantage in treating dry cough due to heat.

[0003] Amino acids are the main active ingredients in asparagus, along with saponins. Amino acids are one of the core material bases for asparagus's traditional effects (nourishing yin and moistening dryness, promoting body fluids and quenching thirst, etc.). Amino acids are hydrophilic and can stimulate the secretion of salivary and sweat glands, relieving symptoms such as dry mouth, sore throat, and dry skin caused by yin deficiency and excessive internal heat or depletion of body fluids (e.g., diabetic dry mouth, menopausal hot flashes and night sweats). Amino acids can also promote the secretion of gastric and intestinal juices, improve digestive enzyme activity, relieve constipation due to yin deficiency (e.g., constipation in the elderly, postpartum constipation), and reduce discomfort caused by dry intestinal mucosa. Therefore, controlling the amino acid content in asparagus is particularly important.

[0004] Currently, the Chinese Pharmacopoeia (2025 edition) for Asparagus lacks amino acid-related characteristic chromatographic testing items. Quality control is limited to thin-layer chromatography identification, moisture content, and extractives, resulting in a one-sided approach that fails to guarantee clinical efficacy. Therefore, comprehensive control is needed. While provincial standards (Shanghai Municipal Standard for Quality of Traditional Chinese Medicine Granules - Asparagus; Beijing Municipal Standard for Quality of Traditional Chinese Medicine Granules - Asparagus) have established characteristic chromatograms for Asparagus, they only target saponins or acid hydrolysates, lacking comprehensive control over amino acid components. Furthermore, the testing methods are too time-consuming (over 30 minutes), making them unsuitable for efficient, batch testing in the industrial production of traditional Chinese medicine. Furthermore, current literature on liquid chromatography detection of asparagus is mostly based on the saponin components of the asparagus medicinal material itself, and basically does not involve other process steps. Therefore, it may not be suitable for the quality detection of asparagus-related preparations. Moreover, the content of saponin components in asparagus is extremely low, the number of peaks is limited, and there are few efficacy-related components. On the other hand, amino acids have more components, more peaks, more information, and are relatively stable and less affected by the process. Therefore, amino acids are more suitable for the quality detection of different asparagus preparations than saponin components.

[0005] Traditional Chinese medicine is primarily used in decoctions, and the amino acid components in Asparagus standard decoctions are one of the important material bases for its efficacy. Therefore, establishing a quality control method based on amino acids is more in line with the characteristics of clinical medication and helps to ensure its effectiveness.

[0006] Traditional Chinese medicine (TCM) formula granules are a new type of prescription medicine made from TCM decoction pieces through processes such as extraction, concentration, drying, and granulation, guided by TCM theory. They have advantages such as convenient administration, easy portability, and controllable quality. However, since Asparagus (Tian Dong) formula granules have lost the morphological identification characteristics of decoction pieces, establishing a comprehensive and rapid quality testing method for Asparagus formula granules is of great significance for achieving overall quality control. Summary of the Invention

[0007] Technical issues

[0008] To address the aforementioned issues, this invention proposes a quality control method applicable to Asparagus and its preparations. This method, based on the characteristics of Asparagus and its preparations (such as Asparagus formula granules and Asparagus standard decoction freeze-dried powder), strengthens specific identification and multi-component, overall quality control, establishes a characteristic spectrum of Asparagus, and conducts comprehensive quality control of Asparagus.

[0009] Technical solution

[0010] The first aspect of this invention provides a method for constructing a quality control method for asparagus and asparagus formulation granules and other asparagus preparations, comprising: (1) preparing a test solution: take 0.2-1 g of the test sample, add 5-20 mL of 10% methanol to dissolve it, sonicate it, take it out, cool it, shake it, and filter it to obtain a test solution; (2) preparing a reference solution; (3) using ultra-high performance liquid chromatography for detection, wherein the conditions of ultra-high performance liquid chromatography include: a chromatographic column with octadecylsilane-bonded silica gel as the packing material; gradient elution with acetonitrile as mobile phase A and 0.1% phosphoric acid solution as mobile phase B, the gradient elution program being: 0-4 min, 1% → 6% A, 99% → 94% B; 4-7 min, 6% → 11%, 94% → 89% B; 7-12 min, 11% → 15%, 89% → 85% B; (4) constructing a characteristic chromatogram and a reference chromatogram of asparagus or its preparations, and comparing them.

[0011] In some embodiments, the ultrasonic treatment in step (1) lasts for 20 to 40 minutes, with a power of 200 to 250 W and a frequency of 40 to 50 kHz.

[0012] In some embodiments, a microporous filter membrane is used for filtration in step (1), wherein the pore size of the microporous filter membrane is 0.22~0.45 μm.

[0013] In some embodiments, the preparation method of the reference solution in step (2) is as follows: take tyrosine, phenylalanine and tryptophan standards, weigh them accurately, add 10% methanol to prepare a solution containing 10 μg of standard per 1 mL, which is the reference solution.

[0014] In some embodiments, the chromatographic column in step (3) has a column length of 150 mm, an inner diameter of 2.1 mm, and a particle size of 1.8 μm.

[0015] In some embodiments, the detection conditions of the ultra-high performance liquid chromatography in step (3) further include: 1) column temperature of 33-37℃; 2) flow rate of 0.38-0.42 mL / min; 3) detection wavelength of 201-205 nm; 4) injection volume of 1-2 μL; 5) the theoretical plate number calculated based on the phenylalanine peak should not be less than 70,000.

[0016] In some embodiments, the quality control method is also applicable to Asparagus medicinal materials or freeze-dried powder of Asparagus standard decoction.

[0017] In some embodiments, the characteristic spectrum includes eight common peaks, wherein peak 4 is a reference peak, peak 2 is tyrosine, peak 4 is phenylalanine, and peak 7 is tryptophan.

[0018] In some embodiments, the relative retention times of the eight common peaks are within ±10% of a specified value, and the specified values ​​for each peak are as follows: peak 1: 0.43, peak 2: 0.52, peak 3: 0.65, peak 4: 1.00, peak 5: 1.18, peak 6: 1.27, peak 7: 1.43, and peak 8: 1.58.

[0019] The second aspect of the present invention provides the application of the construction of the quality control method for any of the above-described asparagus and asparagus formulation granules and other asparagus preparations in the quality control of asparagus and its preparations.

[0020] Technical effect

[0021] 1. In this detection method, the test sample is prepared by ultrasonic treatment with 10% methanol. The preparation method is simple, the operation process is simplified, the method is robust, and it is suitable for different laboratories / production scenarios. At the same time, the amount of organic solvent used is very small, which minimizes environmental pollution.

[0022] 2. In this detection method, the analysis time is only 12 minutes, but the information presented is more comprehensive. Eight common peaks were identified and three characteristic peaks were identified. The baseline is stable and the separation of each peak is good, making it suitable for efficient and batch detection in the industrial production of traditional Chinese medicine.

[0023] 3. The three amino acids identified in this invention are all amino acids containing aromatic ring structures, which can be analyzed and detected with high sensitivity and high repeatability by high-performance liquid chromatography-ultraviolet detection without the need for cumbersome derivatization steps. Other amino acids in aspartic acid with known high content, such as aspartic acid, glutamic acid, and arginine, are aliphatic or basic amino acids, which have virtually no absorption under ultraviolet light and require derivatization for determination. After derivatization, the measured product is not the original product, which may alter the content and polarity of the amino acid, posing a risk of "impurity interference / component destruction".

[0024] 4. This method can be used not only for the quality control of Asparagus granules, but also for the quality control of Asparagus medicinal materials, or freeze-dried powder of Asparagus standard decoction, and other preparations containing Asparagus. Attached Figure Description

[0025] Figure 1 The spectrum contains 8 characteristic peaks; Figure 2 The characteristic spectrum of the asparagus formulation granules in Example 1; Figure 3 The characteristic chromatogram of the freeze-dried powder of Asparagus standard decoction in Example 2; Figure 4 The characteristic spectrum of Asparagus root in Example 3; Figure 5 This is a characteristic peak identification diagram. Detailed Implementation

[0026] To facilitate the implementation of the technical solutions applied for, the terms and expressions involved in this invention will first be explained and defined in general terms and expressions below.

[0027] The terms “comprising,” “including,” or any other variations thereof are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitation, an element defined by the phrase “comprising one…” does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0028] Where specific experimental steps or conditions are not specified in the examples, they can be performed according to the conventional experimental steps or conditions described in the literature in this field. Reagents or instruments whose manufacturers are not specified are all commercially available conventional reagent products. The instruments, reagents, standards, and test samples used in this invention are as follows: Instruments: JY20002 electronic balance (Shanghai Shunyu Hengping Scientific Instruments Co., Ltd.); ME104E electronic balance (Mettler Toledo); XP26 electronic balance (Mettler Toledo Instruments Co., Ltd.); KQ-300DB ultrasonic cleaner (Kunshan Ultrasonic Instruments Co., Ltd.); Waters UPLC ultra-high performance liquid chromatograph.

[0029] Column: Waters ACQUITY UPLC HSS T3 (2.1×150 mm, 1.8 μm).

[0030] Reagents: Acetonitrile (Merk) and phosphoric acid (Fisher Scientific) were chromatographically pure; water was distilled water (Watson's); all other reagents were analytically pure.

[0031] Standards: Tyrosine (batch number: 140609-202516; source: China National Institutes for Food and Drug Control), Phenylalanine (batch number: 140676-202301; source: China National Institutes for Food and Drug Control), Tryptophan (batch number: 140686-202506; source: China National Institutes for Food and Drug Control).

[0032] Test batches: Asparagus granules formula 01, Asparagus granules formula 02, Asparagus granules formula 03, Asparagus granules formula 04, Asparagus granules formula 05; Asparagus standard decoction 01, Asparagus standard decoction 02, Asparagus standard decoction 03, Asparagus standard decoction 04, Asparagus standard decoction 05; Asparagus medicinal materials 01, Asparagus medicinal materials 02, Asparagus medicinal materials 03, Asparagus medicinal materials 04, Asparagus medicinal materials 05.

[0033] Quality control methods include: 1) Preparation of test solution: Take 0.5 g of sample, add 10 mL of 10% methanol, sonicate (250 W, 50 kHz) for 30 min, take out, cool, shake well, filter (0.22 μm) to obtain the test solution.

[0034] 2) Preparation of reference solution: Take appropriate amounts of tyrosine, phenylalanine and tryptophan reference standards, accurately weigh them, add 10% methanol to prepare a solution containing 10 μg per mL, as the reference solution.

[0035] 3) Chromatographic conditions and system suitability

[0036] The column was packed with octadecylsilane-bonded silica gel (150 mm column length, 2.1 mm inner diameter, 1.8 μm particle size); acetonitrile was used as mobile phase A, and 0.1% phosphoric acid solution was used as mobile phase B, with gradient elution performed according to the specifications in Table 1; the detection wavelength was 203 nm; the column temperature was 35 °C; and the flow rate was 0.4 mL / min. The theoretical plate number, calculated based on the phenylalanine peak, should not be less than 70,000.

[0037] Table 1 Gradient elution table

[0038]

[0039] Assay: Accurately pipette 1 μL each of the reference solution and the test solution into the liquid chromatograph, and measure to obtain characteristic chromatograms, such as... Figure 1 As shown.

[0040] The characteristic spectrum shows eight characteristic peaks, of which three peaks (peak 2 tyrosine, peak 4 (S) phenylalanine, and peak 7 tryptophan) should have the same retention time as their corresponding reference peaks. The peak corresponding to the phenylalanine reference peak is the S peak. Calculate the relative retention time and equivalent peak area of ​​the characteristic peak and the S peak. The relative retention time should be within ±10% of the specified value, which is: 0.43 (peak 1), 0.52 (peak 2), 0.65 (peak 3), 1.00 (peak 4S), 1.18 (peak 5), 1.27 (peak 6), 1.43 (peak 7), and 1.58 (peak 8).

[0041] The following provides a further explanation of the construction of the quality control method for asparagus, asparagus formulation granules, and other asparagus preparations provided by this invention.

[0042] Example 1: Detection of Asparagus Formula Granules

[0043] Preparation of test solution: Take 0.5 g of each of 5 batches of Asparagus formulation granules, add 10 mL of 10% methanol, sonicate (250 W, frequency 50 kHz) for 30 min, take out, cool, shake well, filter (0.22 μm) to obtain the test solution.

[0044] Preparation of reference solution: Take appropriate amounts of tyrosine (manufacturer: China National Institute for Food and Drug Control; batch number: 140609-202516), phenylalanine (manufacturer: China National Institute for Food and Drug Control; batch number: 140676-202301), and tryptophan (manufacturer: China National Institute for Food and Drug Control; batch number: 140686-202506) standards, accurately weigh them, and add 10% methanol to prepare a solution containing 10 μg per mL, which is used as the reference solution.

[0045] Chromatographic conditions and system suitability test: Octadecylsilane-bonded silica gel was used as the packing material (column length 150 mm, inner diameter 2.1 mm, particle size 1.8 μm); acetonitrile was used as mobile phase A, and 0.1% phosphoric acid solution was used as mobile phase B, with gradient elution performed according to the specifications in Table 2; the detection wavelength was 203 nm; the column temperature was 35℃; and the flow rate was 0.4 mL / min. The theoretical plate number, calculated based on the phenylalanine peak, should not be less than 70,000.

[0046] Table 2 Gradient Elution Table

[0047]

[0048] Assay: Accurately pipette 1 μL each of the reference solution and the test solution into the liquid chromatograph and determine the result. The determination results are as follows: Figure 2 As shown in Table 3.

[0049] Table 3 Retention time and relative retention time of Asparagus granules

[0050]

[0051] Example 2: Detection of freeze-dried powder of Asparagus standard decoction

[0052] Preparation of test solution: Take 0.5 g of each of 5 batches of freeze-dried Asparagus standard decoction powder, add 10 mL of 10% methanol, sonicate (250 W, frequency 50 kHz) for 30 min, take out, cool, shake well, filter (0.22 μm) to obtain the test solution.

[0053] Preparation of reference solution: Take appropriate amounts of tyrosine (manufacturer: China National Institute for Food and Drug Control; batch number: 140609-202516), phenylalanine (manufacturer: China National Institute for Food and Drug Control; batch number: 140676-202301), and tryptophan (manufacturer: China National Institute for Food and Drug Control; batch number: 140686-202506) standards, accurately weigh them, and add 10% methanol to prepare a solution containing 10 μg per mL, which is used as the reference solution.

[0054] Chromatographic conditions and system suitability test: Octadecylsilane-bonded silica gel was used as the packing material (column length 150 mm, inner diameter 2.1 mm, particle size 1.8 μm); acetonitrile was used as mobile phase A, and 0.1% phosphoric acid solution was used as mobile phase B, with gradient elution performed according to the specifications in Table 4; the detection wavelength was 203 nm; the column temperature was 35℃; and the flow rate was 0.4 mL / min. The theoretical plate number, calculated based on the phenylalanine peak, should not be less than 70,000.

[0055] Table 4 Gradient elution table

[0056]

[0057] Assay: Accurately pipette 1 μL each of the reference solution and the test solution into the liquid chromatograph and determine the result. The determination results are as follows: Figure 3 As shown in Table 5.

[0058] Table 5. Retention time and relative retention time of Asparagus standard decoction

[0059]

[0060] Example 3: Detection of Asparagus Root Powder

[0061] Preparation of the test solution: Take 0.5 g of each of 5 batches of Asparagus powder, add 10 mL of 10% methanol, sonicate (250 W, frequency 50 kHz) for 30 min, take out, cool, shake well, filter (0.22 μm) to obtain the test solution.

[0062] Preparation of reference solution: Take appropriate amounts of tyrosine (manufacturer: China National Institute for Food and Drug Control; batch number: 140609-202516), phenylalanine (manufacturer: China National Institute for Food and Drug Control; batch number: 140676-202301), and tryptophan (manufacturer: China National Institute for Food and Drug Control; batch number: 140686-202506) standards, accurately weigh them, and add 10% methanol to prepare a solution containing 10 μg per mL, which is used as the reference solution.

[0063] Chromatographic conditions and system suitability test: Octadecylsilane-bonded silica gel was used as the packing material (column length 150 mm, inner diameter 2.1 mm, particle size 1.8 μm); acetonitrile was used as mobile phase A, and 0.1% phosphoric acid solution was used as mobile phase B, with gradient elution performed according to the specifications in Table 6; the detection wavelength was 203 nm; the column temperature was 35℃; and the flow rate was 0.4 mL / min. The theoretical plate number, calculated based on the phenylalanine peak, should not be less than 70,000.

[0064] Table 6 Gradient Elution Table

[0065]

[0066] Assay: Accurately pipette 1 μL each of the reference solution and the test solution into the liquid chromatograph and determine the result. The determination results are as follows: Figure 4 As shown in Table 7.

[0067] Table 7. Retention time and relative retention time of Asparagus root.

[0068]

[0069] Experimental Example 1: Methodological Validation

[0070] 1) System Applicability

[0071] Under the selected conditions, a system suitability experiment was conducted using phenylalanine as a reference, and the results are shown in Table 8.

[0072] Table 8 System Suitability Test Results

[0073]

[0074] In summary, the theoretical plate number and repeatability of the phenylalanine peak both meet the analytical requirements. Specifically, the theoretical plate number of the phenylalanine peak should be ≥70,000 to ensure sharp peak shape and baseline separation of the eight characteristic peaks (without interference between peaks). Values ​​below this may result in peak tailing, excessively wide peaks, or other issues affecting separation. Retention time repeatability and peak area repeatability are used to evaluate the precision and stability of the system, ensuring the reliability of the data results.

[0075] 2) Confirmation of characteristic peaks

[0076] Amino acids are the main active components of asparagus. In the study of characteristic chromatograms, the reference solution and the test solution were injected under identical chromatographic conditions. By comparing the chromatograms of the two, peaks in the test sample with "identical retention times" to the reference solution were identified. Ultimately, three characteristic peaks—tyrosine, phenylalanine, and tryptophan—were identified. Figure 5 As shown. The core efficacy of asparagus is "nourishing yin and promoting body fluids, moistening the lungs and clearing the heart," and the three amino acids tyrosine, phenylalanine, and tryptophan are the core material basis for this efficacy. The peak shapes of these three amino acids are stable and well separated, and they can serve as "benchmark peaks" to reflect the overall stability of other components—as long as the relative retention time and relative peak area of ​​these three core peaks meet the standards, it can be inferred that the overall quality of asparagus or the preparation is qualified.

[0077] 3) Repeatability

[0078] Six portions of the asparagus formula granules were taken and their characteristic spectra were obtained according to the method in Example 1. The relative retention time and relative peak area were calculated using peak 4 as the reference peak. The RSD was also calculated. The results are shown in Tables 9 and 10.

[0079] Table 9 Retention Time and Relative Retention Schedule for Asparagus Repeatability Study

[0080]

[0081] Table 10 Peak area and relative peak area of ​​asparagus in repeatability studies

[0082]

[0083] In summary, based on the repeatability test results, the relative retention time RSD of each characteristic peak is in the range of 0.17% to 0.43%, and the relative peak area RSD is in the range of 0.83% to 2.01%, indicating that the repeatability of this characteristic spectrum is good.

[0084] 4) Precision

[0085] One sample of the asparagus formula granules was taken and analyzed according to the method in Example 1. The sample was injected six times consecutively to obtain its characteristic spectrum. Using peak 4 as the reference peak, the relative retention time and relative peak area were calculated. The RSD was also calculated. The results are shown in Tables 11 and 12.

[0086] Table 11 Retention Time and Relative Retention Time for Precision Study of Asparagus

[0087]

[0088] Table 12 Peak Area and Relative Peak Area of ​​Asparagus Precision Examination

[0089]

[0090] In summary, based on the precision test results, the relative retention time RSD of each characteristic peak is in the range of 0.04% to 0.15%, and the relative peak area RSD is in the range of 0.90% to 1.44%, indicating that the instrument has good precision.

[0091] 5) Stability

[0092] One portion of the asparagus formula granules was taken, and a test solution was prepared according to the method in Example 1. The solution was measured at 0 h, 2 h, 4 h, 6 h, 8 h, 10 h, 12 h, and 24 h according to the method in Example 1 to obtain its characteristic spectrum. Using peak 4 as the reference peak, its relative peak area and relative retention time were calculated. The RSD was also calculated, and the results are shown in Tables 13 and 14.

[0093] Table 13 Retention Time and Relative Retention Schedule for Asparagus Stability Study

[0094]

[0095] Table 14 Peak area and relative peak area for stability study of Asparagus

[0096]

[0097] In summary, the stability test results show that the amino acid components in the test solution are stable within 24 hours. Specifically, the relative retention time RSD of each characteristic peak is in the range of 0.12% to 0.61%, and the relative peak area RSD is in the range of 1.22% to 2.69%, indicating that the solution has good stability within 24 hours.

[0098] 6) Intermediate precision

[0099] Using another Shimadzu 30A instrument, 6 portions of the asparagus formula granules were measured according to the method in Example 1 to obtain their characteristic spectra. With peak 4 as the reference peak, their relative retention time was calculated and the RSD was calculated. The results are shown in Table 15.

[0100] Table 15 Retention Time and Relative Retention Time for Intermediate Precision Study of Asparagus

[0101]

[0102] Summary: The data above shows that the relative retention time RSD of each characteristic peak is in the range of 0.04% to 1.22%, and the relative retention time RSD between different instruments is in the range of 0.45% to 2.82%, indicating that the characteristic spectrum has good applicability among different instruments.

[0103] 7) Investigation at different flow velocities

[0104] Take the asparagus formulation granules and prepare the test solution according to the test solution preparation method in Example 1. The test solution was tested at different flow rates of 0.38 mL / min, 0.40 mL / min and 0.42 mL / min according to the method in Example 1 to investigate the robustness of the experimental method to different flow rates. The results are shown in Table 16.

[0105] Table 16. Retention times and relative retention times of each peak at different flow rates

[0106]

[0107] In summary, the data above show that under different flow rates, the relative retention time and relative peak area are both within 5% of the RSD value, indicating that the method has good durability against different flow rates.

[0108] 8) Examination at different wavelengths

[0109] Asparagus granules were taken, and a test solution was prepared according to the method described in Example 1. The solution was then measured at different wavelengths (201 nm, 203 nm, and 205 nm) as described in Example 1. The robustness of the experimental method to different wavelengths was investigated. The results are shown in Table 17.

[0110] Table 17 Retention times and relative retention times of each peak at different wavelengths

[0111]

[0112] Summary: The data above shows that the relative retention time (RSD) is within 5% under different detection wavelengths, indicating that the method has good wavelength robustness.

[0113] 9) Investigation at different column temperatures

[0114] Asparagus granules were taken, and a test solution was prepared according to the method described in Example 1. The solution was then tested at different column temperatures (33℃, 35℃, and 37℃) using the method described in Example 1. The robustness of the experimental method to different column temperatures was investigated. The results are shown in Table 18.

[0115] Table 18. Retention times and relative retention times of each peak at different column temperatures.

[0116]

[0117] In summary, the data above shows that the relative retention time (RSD) is within 5% under different column temperatures, indicating that the method has good durability under column temperature conditions.

[0118] The above specific embodiments further illustrate the purpose, technical solution and beneficial effects of this application. It should be understood that the above are only specific embodiments of this application and are not intended to limit the scope of protection of this application. Any modifications, equivalent substitutions, improvements, etc., made on the basis of the technical solution of this application should be included within the scope of protection of this application.

Claims

1. The construction of a quality control method for asparagus, asparagus formulation granules, and other asparagus preparations, characterized in that, include: (1) Preparation of test solution: Take 0.2-1 g of test sample, add 5-20 mL of 10% methanol to dissolve, sonicate, take out, cool, shake well and filter to obtain test solution; (2) Preparation of reference solution; (3) Detection is performed using ultra-high performance liquid chromatography (UHPLC), wherein the conditions for UHPLC include: A chromatographic column packed with octadecylsilane-bonded silica gel; Gradient elution was performed using acetonitrile as mobile phase A and 0.1% phosphoric acid solution as mobile phase B. The specific gradient elution program was as follows: 0–4 min, 1% → 6% A, 99% → 94% B; 4–7 min, 6% → 11%, 94% → 89% B; 7–12 min, 11% → 15%, 89% → 85% B. (4) Construct characteristic chromatograms and control chromatograms of asparagus or its preparations, and compare them.

2. The construction of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations according to claim 1, characterized in that, The ultrasonic treatment in step (1) lasts for 20 to 40 minutes, with a power of 200 to 250 W and a frequency of 40 to 50 kHz.

3. The construction of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations according to claim 1, characterized in that, In step (1), a microporous filter membrane is used for filtration, and the pore size of the microporous filter membrane is 0.22~0.45 μm.

4. The construction of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations according to claim 1, characterized in that, The method for preparing the reference solution in step (2) is as follows: take tyrosine, phenylalanine and tryptophan standards, weigh them accurately, and add 10% methanol to prepare a solution containing 10 μg of standard per 1 mL, which is the reference solution.

5. The construction of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations according to claim 1, characterized in that, The column in step (3) has a length of 150 mm, an inner diameter of 2.1 mm, and a particle size of 1.8 μm.

6. The construction of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations according to claim 1, characterized in that, The detection conditions for ultra-high performance liquid chromatography described in step (3) also include: 1) Column temperature is 33–37℃; 2) The flow rate is 0.38–0.42 mL / min; 3) The detection wavelength is 201–205 nm; 4) The injection volume is 1–2 μL; 5) The theoretical plate number, calculated based on the phenylalanine peak, should be no less than 70,000.

7. The construction of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations according to claim 1, characterized in that, The test samples include Asparagus root, Asparagus granules, or freeze-dried powder of standard Asparagus decoction.

8. The construction of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations according to claim 1, characterized in that, The characteristic spectrum includes 8 common peaks, of which peak 4 is the reference peak, peak 2 is tyrosine, peak 4 is phenylalanine, and peak 7 is tryptophan.

9. The construction of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations according to claim 8, characterized in that, The relative retention times of the eight common peaks are within ±10% of the specified values, and the specified values ​​for each peak are as follows: Peak 1: 0.43, Peak 2: 0.52, Peak 3: 0.65, Peak 4: 1.00, Peak 5: 1.18, Peak 6: 1.27, Peak 7: 1.43, Peak 8: 1.

58.

10. The application of the quality control method for asparagus and asparagus formulation granules and other asparagus preparations as described in any one of claims 1 to 9 in the quality control of asparagus and its preparations.

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