Construction method of characteristic chromatogram of amomum tsao-ko or amomum tsao-ko kernels or derivative products thereof, and identification method of amomum tsao-ko kernels or derivative products thereof and counterfeit products, amomum tsao-ko with shells or derivative products thereof

Abstract

The invention belongs to the technical field of traditional Chinese medicine detection, and particularly relates to a construction method of a specific chromatogram of amomum tsao-ko or amomum tsao-ko kernels or derivative products thereof and a method for identifying the amomum tsao-ko kernels or the derivative products thereof from counterfeit products, amomum tsao-ko with shells or derivative products thereof. The construction method of the characteristic chromatogram of the amomum tsao-ko or the amomum tsao-ko kernels or the derivative products comprises the step of detecting by adopting a high performance liquid chromatography, and the chromatographic conditions comprise that octadecylsilane chemically bonded silica is used as a filler, a phosphoric acid-containing aqueous solution is used as a mobile phase B, acetonitrile is used as a mobile phase A, and gradient elution is carried out. According to the construction method of the characteristic chromatogram of the amomum tsao-ko or the amomum tsao-ko kernels or the derivative products thereof, the characteristic chromatogram obtained by the method has more characteristic peaks and better baseline separation, water-soluble components and volatile components are synchronously displayed in the same chromatogram, and the method is simple and convenient to operate; therefore, the method is more suitable for quality detection of amomum tsao-ko or amomum tsao-ko kernels or derivative products thereof.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicine detection, and specifically relates to a method for constructing a characteristic spectrum of cardamom or cardamom kernels or their derivatives, and a method for distinguishing cardamom kernels or their derivatives from counterfeits, cardamom with shells or their derivatives. Background Art

[0002] Amomum villosum has multiple pharmacological effects such as gastrointestinal regulation, antibacterial, anti-inflammatory, antioxidant, anti-tumor, hypoglycemic and lipid-regulating. Its main ingredients include volatile oils, phenols, diphenylheptanes and bicyclononanes, etc. When regulating the gastrointestinal system, the active ingredients are volatile oils and polyphenols.

[0003] Amomum tsao-ko Crevost et Lemaire, a plant of the ginger family, is a dried mature fruit. Its seeds are the amomum tsao-ko kernels, which are often used as medicine in traditional Chinese medicine.

[0004] There are studies on the identification of cardamom kernels or their derivatives from common counterfeits in the prior art, but the baseline separation is not good, there are many noise peaks, there is a large slope in the characteristic spectrum and some characteristic peaks are high, which is not conducive to the overall identification and judgment of cardamom, cardamom kernels and their derivatives, and the detection time is long; in addition, when cardamom kernels or their derivatives are used clinically, the cardamom shell in the cardamom needs to be removed and only the cardamom kernels are used, but how to use high performance liquid chromatography to identify cardamom kernels or their derivatives from cardamom with shells is still a blank. How to improve the quality detection method of cardamom or cardamom kernels or their derivatives and effectively control the quality of the finished product is a technical problem that the present invention needs to solve. Summary of the invention

[0005] Therefore, the first object of the present invention is to provide a method for constructing a characteristic spectrum of cardamom or cardamom kernels or their derivatives. The method establishes a characteristic spectrum of the variety according to the characteristics of cardamom or cardamom kernels or their derivatives. The good baseline stability further suppresses the noise peak, controls the overall height balance of the characteristic peak, eliminates the intermediate slope, and facilitates overall identification and judgment, so as to perform comprehensive quality control of cardamom or cardamom kernels or their derivatives.

[0006] The second purpose of the present invention is to provide a method for identifying cardamom and its derivatives or cardamom kernels and its derivatives from counterfeits. The method can identify and distinguish cardamom, cardamom kernels and their derivatives from a variety of common counterfeits and their preparations, including Amomum villosum, Amomum longifolia, Amomum villosum, Amomum villosum, Amomum villosum, Amomum villosum, Amomum villosum, sophora flavescens, wild cardamom, and pseudo-cardamom, so as to better control the quality of cardamom, cardamom kernels and their derivatives.

[0007] The third object of the present invention is to provide a method for identifying cardamom kernels and their derivatives from cardamom with shells and their derivatives. The method can identify and distinguish cardamom kernels and their derivatives from cardamom with shells and their derivatives, facilitate further detection of whether cardamom kernels and their derivatives contain cardamom shells, and facilitate clinical use without using cardamom kernels and their derivatives mixed with cardamom shells.

[0008] To this end, the present invention provides the following technical solutions.

[0009] The present invention provides a method for constructing a characteristic spectrum of tsaoko or tsaoko kernel or a derivative product thereof, comprising detecting by high performance liquid chromatography, wherein the chromatographic conditions include:

[0010] Octadecylsilane bonded silica gel was used as filler, aqueous solution containing phosphoric acid was used as mobile phase B, acetonitrile was used as mobile phase A, and gradient elution was performed. The gradient elution procedure included:

[0011] From 0 to 12 minutes, the volume percentage of mobile phase A was 9%→16%, and the volume percentage of mobile phase B was 91%→84%;

[0012] 12-18 minutes, the volume percentage of mobile phase A is 16%→24%, and the volume percentage of mobile phase B is 84%→76%;

[0013] 18-24 minutes, the volume percentage of mobile phase A is 24%→30%, and the volume percentage of mobile phase B is 76%→70%;

[0014] 24-35 minutes, the volume percentage of mobile phase A is 30%→52%, and the volume percentage of mobile phase B is 70%→48%;

[0015] From 35 to 45 minutes, the volume percentage of mobile phase A is 52%→55%, and the volume percentage of mobile phase B is 48%→45%.

[0016] Optionally, the chromatographic conditions include at least one of the following:

[0017] (1) The detection wavelength is 230-240 nm; optionally, 230 nm for 0 to 30 minutes and 240 nm for 30 to 45 minutes;

[0018] (2) The flow rate is 0.45-0.55 ml / min; optionally, the flow rate is 0.5 ml / min;

[0019] (3) Column temperature 25-35°C; optionally, column temperature 30°C;

[0020] (4) The injection volume is 1-5 μL; optionally, the injection volume is 1-3 μL; further optionally, the injection volume is 2 μL;

[0021] (5) The phosphoric acid concentration in the aqueous solution containing phosphoric acid in the mobile phase B is 0.05 to 0.2% v / v; optionally, the phosphoric acid concentration is 0.1% v / v;

[0022] (6) The specifications of the chromatographic column are: column length 100-150 mm, inner diameter 3.0-4.6 mm, and particle size 1.8-2.7 μm; optionally, the column length is 150 mm, the inner diameter is 3.0 mm, and the particle size is 1.8 μm.

[0023] Optionally, the method further includes the preparation of a test sample solution, including: weighing the test sample, adding a solvent for extraction, separating the solid from the liquid, and taking a liquid, which is the test sample solution.

[0024] Optionally, the preparation of the test solution satisfies at least one of the following:

[0025] A. The ratio of the mass of the test sample to the volume of the solvent is (0.2-1.0):25; the relationship between mass and volume is g / mL;

[0026] B. The extraction method is ultrasonic extraction;

[0027] C. Extraction time is ≥15 minutes, preferably 30 minutes;

[0028] D. The solid-liquid separation is selected from centrifugation or membrane filtration;

[0029] E. The solvent is selected from one or more of methanol, ethanol and water; preferably, it is a methanol-water solution with a volume percentage of 50-70%.

[0030] Optionally, the construction method further comprises the steps of preparing a reference solution using at least one of catechin, epicatechin and citral, and obtaining a reference spectrum of the reference solution by detecting the reference solution using the high performance liquid chromatography method in the construction method;

[0031] Preferably, each 1 mL of catechin reference solution contains 10-30 μg of catechins; preferably, 20 μg;

[0032] Preferably, each 1 mL of epicatechin reference solution contains 20-40 μg of epicatechin; preferably, 30 μg;

[0033] Preferably, each 1 mL of citral reference solution contains 20-40 μg of citral; preferably, 30 μg;

[0034] Preferably, the solvent used in the preparation of the reference solution is selected from methanol or a methanol aqueous solution with a volume fraction of not less than 50-70%; more preferably, the solvent used in the preparation of the reference solution is selected from a 70% methanol aqueous solution;

[0035] Preferably, the construction method further comprises the step of preparing a control medicinal material solution using the Amomum villosum control medicinal material, and the step of detecting the control medicinal material solution by the high performance liquid chromatography method in the construction method to obtain a control medicinal material reference atlas;

[0036] Preferably, the preparation method of the control medicinal material solution is the same as the preparation method of the test solution;

[0037] Preferably, the amomum villosum or its derivative products include at least one of amomum villosum medicinal materials, amomum villosum decoction pieces or amomum villosum preparations; the amomum villosum kernels or their derivative products include at least one of amomum villosum kernel decoction pieces or amomum villosum preparations.

[0038] The present invention also provides a method for determining the content of tsaoko or tsaoko kernel or its derivative products, comprising:

[0039] Take the test solution and the reference solution, and respectively detect them by the high performance liquid chromatography method in the method for constructing the characteristic spectrum of the tsaoko or tsaoko kernel or its derivative products;

[0040] Wherein, the reference substance includes at least one of catechin, epicatechin and citral.

[0041] The present invention also provides a quality detection method for the tsaoko or tsaoko kernel or a product derived therefrom, comprising the steps of comparing a characteristic spectrum of the product to be detected with a control characteristic spectrum of the tsaoko or tsaoko kernel or a product derived therefrom;

[0042] The characteristic spectrum of the product to be tested is constructed according to the construction method;

[0043] The reference characteristic spectrum of the Amomum villosum or Amomum villosum kernel or its derivative products is selected from any one of the following (1)-(4):

[0044] (1) It has 10 common characteristic peaks. The peak corresponding to the reference peak of epicatechin is the S peak. The relative retention times of peaks 1-2, 4, 6-8 and S peak are calculated. The relative retention times should be within the range of ±10% of the specified values. The specified values ​​of peaks 1-2, 4 and 6-8 are 0.41, 0.61, 0.87, 1.15, 1.31 and 1.78 respectively.

[0045] (2) It has 10 common characteristic peaks, of which 4 peaks correspond to the retention times of reference peaks of catechin, epicatechin and citral respectively, and the peak corresponding to the reference peak of epicatechin is the S peak. The relative retention times of peaks 1-2, peak 4 and peaks 6-8 and the S peak are calculated, and the relative retention times should be within the range of ±10% of the specified values. The specified values ​​of peaks 1-2, peak 4 and peak 6-8 are: 0.41, 0.61, 0.87, 1.15, 1.31, 1.78 respectively; optionally, peak 1 corresponds to protocatechuic acid; peak 2 corresponds to protocatechuic aldehyde; peak 3 corresponds to catechin; peak 4 corresponds to proanthocyanidin B2; peak 5 (S) corresponds to epicatechin; peaks 9 and 10 correspond to citral;

[0046] (3) Using a single batch or multiple batches of cardamom and / or cardamom kernels and / or their derivatives as test samples, the characteristic spectrum of cardamom kernels and / or their derivatives obtained according to the construction method described above;

[0047] (4) Using multiple batches of cardamom and / or cardamom kernels and / or their derivatives as test samples, the characteristic spectra obtained according to the construction method are prepared into a control characteristic spectrum by using the average value or median method.

[0048] The present invention also provides a method for distinguishing between tsaoko or tsaoko kernels or derivative products thereof and counterfeits, comprising:

[0049] The test is carried out in accordance with the above-mentioned quality test method for the tsaoko or tsaoko kernel or its derivatives; wherein, for the tsaoko or tsaoko kernel or its derivatives, the relative peak area of ​​peak 7 and peak S shall be within the prescribed range, and the prescribed value shall be: not less than 0.31;

[0050] Optionally, the counterfeit products include one or more of Amomum villosum or its derivatives, Alpinia cyrtonema or its derivatives, Amomum villosum or its derivatives, Amomum villosum or its derivatives; preferably, the Amomum villosum includes one or more of Amomum villosum yangchunensis, Amomum villosum longifolia, Amomum villosum niuginosa, and Amomum villosum fragrant.

[0051] The present invention also provides a method for identifying tsaoko kernels or derivatives thereof and tsaoko in shell or derivatives thereof, comprising:

[0052] The product to be tested is subjected to a step of detecting a characteristic spectrum by high performance liquid chromatography in the construction method;

[0053] The chromatographic conditions of the high performance liquid chromatography method include a detection wavelength of 192-194 nm;

[0054] The reference solution also includes a reference solution containing 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol;

[0055] For the tsaoko kernel or its derivative products, the peak area of ​​the chromatographic peak of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in the characteristic spectrum is ≤ the peak area of ​​the chromatographic peak of the corresponding reference solution of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol;

[0056] For shelled amomum villosum or its derivative products, the chromatographic peak area of ​​1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in the characteristic spectrum is greater than the chromatographic peak area of ​​the corresponding reference solution of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol.

[0057] Optionally, each 1 mL of the reference solution contains 1.6-2.5 μg of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol or 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol;

[0058] Preferably, for shelled cardamom or its derivative products, the chromatographic peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol is detected to be greater than 155239 μAU·S; for cardamom kernel or its derivative products, the chromatographic peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol is detected to be ≤155239 μAU·S.

[0059] The technical solution of the present invention has the following advantages:

[0060] 1. The method for constructing a characteristic spectrum of tsaoko or tsaoko kernel or its derivative products provided by the present invention uses octadecylsilane bonded silica gel as a filler, and the mobile phase includes an aqueous solution containing phosphoric acid and acetonitrile. Through a specific gradient elution program, 10 common characteristic peaks are obtained, and effective separation of the common characteristic peaks is achieved. Moreover, the obtained characteristic spectrum has more characteristic peaks and better baseline separation, and the water-soluble components and volatile components are simultaneously displayed in the same chromatogram. The method is simple, easy to operate, and has a short detection time. The peak positions of protocatechuic acid, protocatechuic aldehyde, proanthocyanidin B2, catechin, protocatechin, and citral can be accurately located, fully reflecting the integrity and characteristic of tsaoko or tsaoko kernel or its derivative products (such as tsaoko kernel formula granules and other derivative products), and providing a basis for quality detection and content determination of tsaoko or tsaoko kernel or its derivative products.

[0061] 2. The method for constructing the characteristic spectrum of the cardamom or cardamom kernel or its derivative products provided by the present invention is to determine the optimal chromatographic conditions by optimizing the chromatographic conditions, so that the peak area is higher and the separation effect is better, so that the quality of the cardamom, cardamom kernel or its derivative products can be more comprehensively monitored.

[0062] 3. The method for identifying the cardamom or cardamom kernels or their derivatives from counterfeits provided by the present invention can effectively identify the cardamom or cardamom kernels or their derivatives from counterfeits by comparing the characteristic spectrum of the product to be identified with the control characteristic spectrum of the cardamom or cardamom kernels or their derivatives. The counterfeits include Amomum villosum and its preparations, Amomum cardamomum seeds and its preparations, wild cardamom and its preparations, and Pseudo-cardamom and its preparations; the Amomum villosum includes Amomum villosum, Amomum villosum longifolia, Amomum villosum niuginosa, and Amomum villosum fragrant.

[0063] 4. The method for distinguishing between the grass kernel or its derivative products and the grass kernel with shell or its derivative products provided by the present invention comprises the following steps: by detecting whether the product to be tested contains 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol, a meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference solution is prepared; and by comparing the peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference solution with a concentration of 1.6-2.5 μg / mL with the peak area of ​​meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in the product to be tested, it is possible to effectively detect whether the product to be tested contains grass shell, thereby ensuring that the grass kernel or its derivative products mixed with grass shell will not be used in clinical use.

[0064] 5. The quality detection method of the cardamom or cardamom kernel or its derivative products provided by the present invention compares the characteristic spectrum of the cardamom or cardamom kernel product to be tested with the control characteristic spectrum of the cardamom or cardamom kernel or its derivative products, so as to more comprehensively detect the quality of the cardamom or cardamom kernel or its derivative products, thereby proving that the quality of the product is effective and controllable. BRIEF DESCRIPTION OF THE DRAWINGS

[0065] In order to more clearly illustrate the specific implementation methods of the present invention or the technical solutions in the prior art, the drawings required for use in the specific implementation methods or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are some implementation methods of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0066] FIG1 is a characteristic spectrum of the tsaoko kernel formula granules; specifically, FIG1-(1) is the test sample; FIG1-(2) is catechin; FIG1-(3) is epicatechin; FIG1-(4) is citral; FIG1-(5) is the tsaoko control medicinal material; FIG1-(6) is the tsaoko kernel control medicinal material;

[0067] Figure 2 is the chromatogram of gradient condition 1 in Experimental Example 1;

[0068] Figure 3 is the chromatogram of gradient condition 2 in Experimental Example 1;

[0069] Figure 4 is the chromatogram of gradient condition 3 in Experimental Example 1;

[0070] Figure 5 is the chromatogram of gradient condition 4 in Experimental Example 1;

[0071] Figure 6 is the chromatogram of gradient condition 5 in Experimental Example 1;

[0072] Figure 7 is the chromatogram of gradient condition 6 in Experimental Example 1;

[0073] Figure 8 is the chromatogram of gradient condition 7 in Experimental Example 1;

[0074] Fig. 9 is the chromatogram of gradient condition 8 in Experimental Example 1;

[0075] Fig.10 is the chromatogram of gradient condition 9 in Experimental Example 1;

[0076] Fig.11 is the chromatogram of gradient condition 10 in Experimental Example 1;

[0077] Fig.12 This is the chromatogram of the flow rate of 0.45 ml / min in the flow rate investigation in Experimental Example 1;

[0078] Fig.13 This is the chromatogram of the flow rate of 0.50 ml / min in the flow rate investigation in Experimental Example 1;

[0079] Fig.14 This is the chromatogram of the flow rate of 0.55 ml / min in the flow rate investigation in Experimental Example 1;

[0080] Fig.15 This is the chromatogram with the column temperature at 25°C in the investigation of column temperature in Experimental Example 1;

[0081] Fig.16 This is the chromatogram with the column temperature at 30°C in the investigation of column temperature in Experimental Example 1;

[0082] Fig.17 This is the chromatogram with the column temperature at 35°C in the investigation of column temperature in Experimental Example 1;

[0083] Fig.18This is a chromatogram of the mobile phase of acetonitrile-water in the mobile phase investigation in Experimental Example 1;

[0084] Fig.19 This is a chromatogram of the mobile phase of acetonitrile-0.1% formic acid aqueous solution in the mobile phase investigation in Experimental Example 1;

[0085] Fig. 20 The chromatogram of the mobile phase in the investigation of the mobile phase in Experimental Example 1 is acetonitrile-0.1% glacial acetic acid aqueous solution;

[0086] Fig.21 The chromatogram of the mobile phase in the investigation of the mobile phase in Experimental Example 1 is acetonitrile-0.05% phosphoric acid aqueous solution;

[0087] Fig. 22 This is a chromatogram of the mobile phase of acetonitrile-0.1% phosphoric acid aqueous solution in the mobile phase investigation in Experimental Example 1;

[0088] Fig.23 This is a chromatogram of the mobile phase of acetonitrile-0.2% phosphoric acid aqueous solution in the mobile phase investigation in Experimental Example 1;

[0089] Fig.24 This is a chromatogram with a detection wavelength of 230 nm in the investigation of the detection wavelength in Experimental Example 1;

[0090] Fig.25 This is a chromatogram with a detection wavelength of 240 nm in the investigation of the detection wavelength in Experimental Example 1;

[0091] Fig.26 This is a chromatogram with a detection wavelength of 254 nm in the investigation of the detection wavelength in Experimental Example 1;

[0092] Fig. 27 The chromatogram is a chromatogram in which the detection wavelength is 230 nm for 0-30 minutes and 240 nm for 30-45 minutes in the investigation of the detection wavelength in Experimental Example 1;

[0093] Fig.28 This is the chromatogram of the injection volume of 1 μL in the injection volume investigation in Experimental Example 1;

[0094] Fig.29 This is the chromatogram of the injection volume of 2 μL in the injection volume investigation in Experimental Example 1;

[0095] Fig.30 This is the chromatogram of the injection volume of 3 μL in the injection volume investigation in Experimental Example 1;

[0096] Fig.31 This is the chromatogram of the injection volume of 4 μL in the injection volume investigation in Experimental Example 1;

[0097] Fig.32 This is the chromatogram of the injection volume of 5 μL in the injection volume investigation in Experimental Example 1;

[0098] Fig.33 Characteristic spectra of three batches of grass kernel formula granules in Experimental Example 2;

[0099] Fig.34 This is the characteristic spectrum of the reference medicinal material Amomum villosum in Experimental Example 3;

[0100] Fig.35 It is the characteristic spectrum of 17 batches of standard decoction (lyophilized powder) of Tsaoko kernel slices in Experimental Example 3 (the batch numbers from bottom to top are: 2103001Y, 2103002Y, 2103003Y, 2103004Y, 2103005Y, 2103006Y, 2103007Y, 2103008Y, 2103009Y, 2103010Y, 2103011Y, 2103013Y, 2103014Y, 2103015Y, 230301Y, 230302Y, 230303Y);

[0101] Fig.36 is the reference characteristic spectrum; among them, peak 1: protocatechuic acid; peak 2: protocatechuic aldehyde; peak 3: catechin; peak 4: proanthocyanidin B2; peak 5: epicatechin; peak 9 and peak 10: citral;

[0102] Fig.37 This is the reference sample positioning map; from bottom to top in the figure: Tsaoko kernel formula granules (batch number: 2303001Y); protocatechuic acid reference; protocatechuic aldehyde reference; catechin reference; proanthocyanidin B2 reference; epicatechin reference; citral reference;

[0103] Fig.38 It is the chromatogram of negative blank solution;

[0104] Fig.39 Comparison of the characteristic spectra of Amomum villosum kernel and Amomum villosum pseudo-Amomum villosum; S1-3: Amomum villosum pseudo-Amomum villosum; S4: Amomum villosum kernel;

[0105] Fig.40 Comparison of the characteristic spectra of the tsaoko kernel and the wild tsaoko fruit; S1-3: wild tsaoko fruit; S4: tsaoko kernel;

[0106] Fig.41 The characteristic spectrum of Amomum villosum kernel is compared with that of Amomum villosum and Amomum cyrtonema; S1-3: Amomum villosum; S4: Amomum villosum; S5: Amomum villosum; S6: Amomum villosum; S7: Amomum cyrtonema; S8: Amomum villosum kernel;

[0107] Fig.42 This is the UV spectrum of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol;

[0108] Fig.43This is the UV spectrum of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol;

[0109] Fig.44 These are the chromatograms of samples related to Amomum villosum and Amomum villosum kernels at a wavelength of 192 nm; S1: Amomum villosum medicinal material; S2: Amomum villosum kernel decoction pieces; S3: Amomum villosum freeze-dried powder; S4: Amomum villosum kernel freeze-dried powder; S5: 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substance; S6: meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substance. DETAILED DESCRIPTION

[0110] The following examples are provided for a better understanding of the present invention, but are not intended to limit the best mode of implementation, nor to limit the content and protection scope of the present invention. Any product identical or similar to the present invention obtained by anyone under the inspiration of the present invention or by combining the features of the present invention with other prior arts shall fall within the protection scope of the present invention.

[0111] If no specific experimental steps or conditions are specified in the examples, the conventional experimental steps or conditions described in the literature in the field can be used. If no manufacturer is specified for the reagents or instruments used, they are all conventional reagent products that can be obtained commercially.

[0112] The test sample in the following embodiments and experimental examples is selected from the formula granules of grass kernels. The specific preparation method of the formula granules of grass kernels is as follows: 6800g of grass kernel slices are taken, crushed, boiled and extracted, 18 times the mass of water is added, extracted for 1.5 hours, and the extract is obtained while collecting a sufficient amount of volatile oil. The volatile oil is taken by grinding method, 7 times the mass of volatile oil is added to prepare inclusion compound, the extract is filtered through a 200 mesh filter cloth while hot, the filtrate is concentrated under reduced pressure at 70°C to a relative density of 1.03 to 1.10 (65±5°C), the inclusion compound is added to the concentrate, mixed, spray dried, crushed, dry granulated, packaged, and sealed for storage.

[0113] The standard decoction of the tsaoko kernel slices of the present invention is prepared by the following method: taking the tsaoko kernel slices, breaking them, adding 2 times the amount of water to soak for 30 minutes, adding 6 times the amount of water, boiling with boiling water and slow fire (200W) for 5 minutes, filtering with a 200-mesh filter cloth while hot, adding 6 times the amount of water to the residue, boiling with high fire (500W), then turning to slow fire (200W) and boiling for 25 minutes, filtering with a 200-mesh filter cloth while hot, combining the two filtrates, and the filtrate is the standard decoction of the tsaoko kernel slices.

[0114] The main instruments and reagents involved in the present invention are as follows:

[0115] 1. Instruments, reagents and test drugs

[0116] Chromatograph 1: Waters ACQUITY UPLC H-Class PLUS chromatography system, including quaternary solvent manager (ACQ-QSM), sample manager (ACQ-FTN), imported column oven (ACQ-CM), diode array UV detector (ACQ-PDA), and Empower chromatography management system;

[0117] Chromatograph 2: Agilent 1290 Infinity II chromatography system, including quaternary solvent manager (1290 Flexible Pump), sample manager (1290 Vialsampler), original imported chromatography column oven (1290 MCT), diode array UV detector (1290 PDA FS), OpenLAB CDS chromatography management system;

[0118] Chromatograph 3: Thermo Vanquish chromatography system, including quaternary solvent manager (Pump), sample manager (Autosampler), original imported chromatography column oven (Column-Compartment), diode array ultraviolet detector (Detector), and Chromeleon 7.2 chromatography management system.

[0119] Chromatographic column: Waters HSST3 (column length is 150 mm, inner diameter is 3.0 mm, and particle size is 1.8 μm); CAPCELLCOREC18 (column length is 150 mm, inner diameter is 4.6 mm, and particle size is 2.7 μm); Welch core-shell (column length is 150 mm, inner diameter is 4.6 mm, and particle size is 2.7 μm).

[0120] Ultrasonic instrument: KQ-500DE (Kunshan Ultrasonic Instrument Co., Ltd.)

[0121] 2. Reagents and test drugs:

[0122] Protocatechuic acid reference substance (batch number: 110809-202207, purchased from China Food and Drug Inspection Institute, purity 97.5%); protocatechuic aldehyde reference substance (batch number: 110810-202210, purchased from China Food and Drug Inspection Institute, purity 99.9%); citral reference substance (batch number: 190178-201701, purchased from China Food and Drug Inspection Institute); catechin reference substance (batch number: 110877-202005, purchased from China Food and Drug Inspection Institute, purity 95.1%); epicatechin reference substance (batch number: 110878-201703, purchased from China Food and Drug Administration, purity 99.7%); proanthocyanidin B2 reference substance (batch number: 000897-202006, purchased from Jiangxi Baicaoyuan Biotechnology Co., Ltd.); Amomum villosum reference medicinal material (batch number: 121550-201602, purchased from China Food and Drug Administration); Amomum villosum kernel reference medicinal material (batch number: 230138-202401, Shanghai Hongyong Biotechnology Co., Ltd.). Acetonitrile was chromatographically pure (Thermo Fisher Scientific Inc.), water was ultrapure water, and other reagents were analytically pure.

[0123] Grass kernel formula granules (batch numbers: 2303001Y, 2303002Y, 2303003Y).

[0124] As used herein, the grass fruit refers to the grass fruit shell and the grass fruit kernel, which is the same as the grass fruit with shell.

[0125] Example 1

[0126] This embodiment provides a method for constructing a characteristic spectrum of tsaoko or tsaoko kernel or its derivative products, comprising:

[0127] (1) Preparation of test solution: Take the tsaoko kernel formula granules as the test sample, take about 0.2 g of the test sample powder, accurately weigh it, put it into a stoppered conical flask, add 25 ml of 70% v / v methanol aqueous solution, weigh it, and ultrasonically treat it (power 250 W, frequency 40 kHz) for 30 minutes, take it out, cool it, make up the lost weight with 70% v / v methanol aqueous solution, shake it well, filter it, and take the filtrate to obtain the solution;

[0128] Preparation of reference medicinal material solution: Take 1g of each of the Amomum villosum reference medicinal material and the Amomum villosum kernel reference medicinal material, and prepare the Amomum villosum reference medicinal material solution and the Amomum villosum kernel reference medicinal material solution according to the above "Preparation of Test Solution";

[0129] Preparation of reference substance solution: Take appropriate amount of epicatechin and citral reference substances, weigh them accurately, add methanol to make a solution containing 30μg per 1ml, and prepare epicatechin reference substance solution and citral reference substance solution. Take appropriate amount of catechin reference substance, weigh them accurately, add methanol to make a solution containing 20μg per 1ml, and prepare catechin reference substance solution.

[0130] (2) Take 2 μL of the test solution, the reference medicinal material solution and the reference substance solution and use high performance liquid chromatography to measure the content of the sample. The chromatographic conditions are as follows: octadecylsilane bonded silica gel is used as the filler (Waters HSST3) column length 150mm, inner diameter 3.0mm, particle size 1.8μm); acetonitrile as mobile phase A, 0.1% (v / v) phosphoric acid solution as mobile phase B, gradient elution as specified in the following table; flow rate 0.5ml per minute; column temperature 30℃; detection wavelength 230nm for 0-30 minutes, 240nm for 30-45 minutes. The theoretical plate number calculated based on the epicatechin peak should be no less than 5000.

[0131] Table 1 Gradient elution table

[0132]

[0133] The results are shown in Table 2 and Figure 1. Figure 1-(1) to Figure 1-(6) It can be seen that the characteristic spectrum of the tsaoko kernel formula granules corresponds to the chromatographic peak of the characteristic spectrum of the reference medicinal material. The characteristic spectrum of the tsaoko kernel formula granules has 10 characteristic peaks, and all 10 characteristic peaks have been effectively separated. Each characteristic peak has a good peak shape, a stable baseline, a short detection time, and a uniform peak height or peak area. Among them, peak 3 and peak 5 correspond to the retention time of the catechin reference substance peak and the epicatechin reference substance peak, respectively, and peak 9 and peak 10 correspond to the retention time of the citral reference substance peak; the peak corresponding to the retention time of the epicatechin reference substance peak is the S peak, and the relative retention times of peaks 1-2, peak 4 and peaks 6-8 and the S peak are calculated, and their relative retention times are within the range of ±10% of the specified value, and the specified values ​​are: 0.41 (peak 1), 0.61 (peak 2), 0.87 (peak 4), 1.15 (peak 6), 1.31 (peak 7), and 1.78 (peak 8). Among them, peak 1 corresponds to protocatechuic acid; peak 2 corresponds to protocatechuic aldehyde; peak 3 corresponds to catechin; peak 4 corresponds to proanthocyanidin B2; peak 5 (S) corresponds to epicatechin; peaks 9 and 10 correspond to citral (peaks 9 and 10 correspond to the two isomers of citral, neral and geranial).

[0134] Both the cardamom fruit control medicinal material and the cardamom kernel control medicinal material show 10 characteristic peaks, and the chromatographic peaks are not much different. Considering that the current control supplied by the China National Institute for Food and Drug Control is the cardamom fruit control medicinal material, and the cardamom kernel control medicinal material is provided by a third-party reference material company, the supply quality of the cardamom fruit control medicinal material is more guaranteed. Therefore, the cardamom fruit control medicinal material is used as the control, and the cardamom kernel control medicinal material is no longer used as the control.

[0135] Table 2 System suitability results

[0136]

[0137]

[0138] Experimental Example 1

[0139] 1. Investigation of chromatographic conditions

[0140] 1. Selection of mobile phase gradient

[0141] Take the same sample solution prepared in Example 1, and inject 2 μL of it; use octadecylsilane bonded silica gel as the filler (Waters HSST3) column length 150mm, inner diameter 3.0mm, particle size 1.8μm); acetonitrile as mobile phase A, 0.1% phosphoric acid solution (V / V) as mobile phase B, gradient elution according to the provisions in Table 3; flow rate 0.3ml per minute; column temperature 30℃; detection wavelength 240nm, the results are shown in Figure 2 .

[0142] Table 3 Gradient conditions 1

[0143]

[0144] Depend on Figure 2 It can be seen that the gradient elution was performed using gradient condition 1. The water-soluble components with greater polarity were eluted in 0-20 minutes, and the non-polar components of volatile oils were eluted in 30-40 minutes. There was basically no chromatographic peak after 40 minutes. However, the chromatographic peak separation was poor. Therefore, the mobile phase ratio was further optimized and gradient elution was performed according to the regulations in Table 4. The results are shown in Figure 3 ;

[0145] Table 4 Gradient conditions 2

[0146]

[0147] Depend on Figure 3 It can be seen that the elution using gradient condition 2 has a poor separation effect on the chromatographic peak of water-soluble components in 10-20 minutes. Further optimization was performed and gradient elution was performed according to the provisions in Table 5. The results are shown in Figure 4 ;

[0148] Table 5 Gradient conditions 3

[0149]

[0150] Depend on Figure 4 It can be seen that the separation of each chromatographic peak is improved by using gradient condition 3 for elution, but the analysis time is longer. Further optimization is carried out according to the provisions in Table 6 for gradient elution. The results are shown in Figure 5 ;

[0151] Table 6 Gradient Condition 4

[0152]

[0153] Depend on Figure 5 It can be seen that the analysis time becomes shorter when gradient condition 4 is used for elution and the elution flow rate and elution gradient are adjusted, but the separation of the chromatographic peaks from 0 to 7 minutes is poor. Further optimization is performed and gradient elution is performed according to the provisions in Table 7. The results are shown in FIG. Figure 6 ;

[0154] Table 7 Gradient Condition 5

[0155]

[0156]

[0157] Depend on Figure 6 It can be seen that the elution was performed using gradient condition 5, and the chromatographic peaks were well separated in the period of 0-20 minutes, but the baseline was not stable due to the rapid gradient change in the period of 20-30 minutes. Further optimization was performed and the gradient elution was performed according to the provisions in Table 8. The results are shown in Table 8. Figure 7 ;

[0158] Table 8 Gradient Condition 6

[0159]

[0160] Depend on Figure 7 It can be seen that the problem of unstable baseline can be improved by slowing down the gradient change when using gradient condition 6 for elution, but the analysis time is increased. Further optimization is performed according to the provisions in Table 9 for gradient elution. The results are shown in Figure 8 ;

[0161] Table 9 Gradient Conditions 7

[0162]

[0163] Depend on Figure 8 It can be seen that the chromatographic peak separation at 6-10 minutes is poor when gradient condition 7 is used for elution. The elution gradient is adjusted and the analysis time is shortened. Further optimization is performed and gradient elution is performed according to the provisions in Table 10. The results are shown in Fig. 9 ;

[0164] Table 10 Gradient Conditions 8

[0165]

[0166]

[0167] Depend on Fig. 9 It can be seen that the separation effect of each characteristic peak is good when the gradient condition 8 is used for elution. The elution gradient is slightly adjusted to further shorten the analysis time. The gradient elution is performed according to the provisions in Table 11. The results are shown in Fig.10 ;

[0168] Table 11 Gradient conditions 9

[0169]

[0170] Depend on Fig.10 It can be seen that when gradient condition 9 is used for elution, the chromatographic peak separation is poor due to the rapid change of the elution gradient in 13-25 minutes. Further optimization is carried out according to the provisions in Table 12. The results are shown in Fig.11 ;

[0171] Table 12 Gradient conditions 10

[0172]

[0173] Depend on Fig.11 It can be seen that when gradient condition 10 is used for elution, the separation effect of each characteristic peak is better and the analysis time is shorter. Therefore, gradient condition 10 is temporarily set as the elution gradient, and the flow rate, column temperature and detection wavelength are further investigated.

[0174] 2. Investigation of flow rate

[0175] In addition to the flow rate, the chromatographic conditions determined by the selection of the mobile phase gradient in 1 were used, with the flow rate as the variable. The same portion of the test solution prepared in Example 1 was taken and measured at flow rates of 0.45 ml / min, 0.50 ml / min, and 0.55 ml / min, respectively. The resulting chromatograms are shown in FIG. Figure 12-14 As shown. Figure 12-14 It can be seen that when the elution flow rate changes slightly, the separation differences of the characteristic peaks are not obvious, so the flow rate is temporarily set at 0.50 ml / min for subsequent condition screening.

[0176] 3. Investigation of column temperature

[0177] In addition to the column temperature, the chromatographic conditions determined by the investigation of 2. Flow rate, with column temperature as the variable, took the same portion of the test solution prepared according to Example 1, and measured it at column temperatures of 25°C, 30°C, and 35°C, respectively. The resulting chromatograms are shown in Figure 15-17 As shown. Figure 15-17It can be seen that the change of column temperature has a great influence on the separation of the chromatographic peaks in 10-25 minutes. The chromatographic effect obtained by a column temperature of 30°C is relatively better. Therefore, the column temperature is temporarily set at 30°C for subsequent condition screening.

[0178] 4. Investigation of mobile phase

[0179] In addition to the mobile phase, according to the chromatographic conditions determined by the investigation of 3. Column temperature, with the mobile phase as the variable, the same portion of the test solution prepared according to Example 1 was taken and measured under different mobile phases (acetonitrile-water, acetonitrile-0.1% v / v formic acid aqueous solution, acetonitrile-0.1% v / v glacial acetic acid aqueous solution, acetonitrile-0.05% v / v phosphoric acid aqueous solution, acetonitrile-0.1% v / v phosphoric acid aqueous solution, acetonitrile-0.2% v / v phosphoric acid aqueous solution), and the obtained chromatograms are shown in FIG. Figure 18-23 As shown. Figure 18-23 It can be seen that different mobile phases have a great influence on the characteristic spectra. When formic acid or glacial acetic acid is selected as the mobile phase, the baseline is not stable. When 0.1% phosphoric acid aqueous solution is used as the mobile phase, the chromatographic peak separation and peak shape are better. Therefore, the mobile phase is temporarily set as acetonitrile-0.1% phosphoric acid aqueous solution for subsequent condition screening.

[0180] 5. Investigation of detection wavelength

[0181] In addition to the detection wavelength, according to the chromatographic conditions determined in 4. Investigation of the mobile phase, with the detection wavelength as a variable, the same sample solution prepared according to Example 1 was taken and measured at different detection wavelengths of 230 nm, 240 nm, 254 nm (230 nm for 0-30 minutes and 240 nm for 30-45 minutes). The resulting chromatogram is shown in FIG. Figure 24-27 As shown. Figure 24-27 It can be seen that different detection wavelengths have a great influence on the chromatographic peak response. At a wavelength of 230nm, the chromatographic peak response of the water-soluble components in the first half of the characteristic spectrum is better. At a wavelength of 240nm, the chromatographic peak response of the volatile components in the second half of the characteristic spectrum is better. At a wavelength of 254nm, the overall chromatographic peak response is lower. Therefore, it is chosen to switch the wavelength from 230nm to 240nm in 30 minutes to ensure that there is enough time to switch the wavelength without affecting the detection of volatile oil components. Therefore, the detection wavelength is temporarily set at 230nm from 0 to 30 minutes and 240nm from 30 to 45 minutes for subsequent condition screening.

[0182] 6. Sample volume inspection

[0183] According to the chromatographic conditions determined in 5. Detection wavelength, with the injection volume as the variable, take the same portion of the test solution prepared according to Example 1, and accurately aspirate 1 μL, 2 μL, 3 μL, 4 μL, and 5 μL of the test solution for measurement. The resulting chromatogram is shown in Figure 28-32 As shown. Figure 28-32It can be seen that different injection volumes have a greater impact on the chromatographic peak response. When the injection volume is 4μL or 5μL, the chromatographic peak is deformed. When the injection volume is 2μL, the chromatographic peak shape is better and the peak response is moderate. Therefore, the injection volume is tentatively set to 2μL.

[0184] 7. Determination of optimal chromatographic conditions

[0185] Octadecylsilane bonded silica gel as filler (Waters HSST3) column length 150 mm, inner diameter 3.0 mm, particle size 1.8 μm); acetonitrile as mobile phase A, 0.1% (v / v) phosphoric acid solution as mobile phase B, gradient elution according to the provisions in Table 13; flow rate 0.5 ml per minute; column temperature 30°C; detection wavelength 230 nm for 0-30 minutes, 240 nm for 30-45 minutes. Injection volume 2 μL.

[0186] Table 13 Gradient elution table

[0187]

[0188] Experimental Example 2 Construction of characteristic spectra of three batches of cardamom kernel formula granules

[0189] Three batches of cardamom kernel formula granule samples were taken as test samples, and test sample solutions were prepared according to the method in Example 1; cardamom control medicinal materials were taken according to the method in Example 1 to prepare cardamom control medicinal material reference solution; catechin, epicatechin, and citral reference substances were taken according to the method in Example 1 to prepare catechin reference substance reference solution, epicatechin reference substance reference solution, and citral reference substance reference solution.

[0190] The above solution was detected by high performance liquid chromatography of Example 1 to obtain characteristic spectra of three batches of grass kernel formula particles. The results are shown in Tables 14-15 (Note: in Table 14, since the switching wavelength is 240nm at 30-45 minutes, peak 9 is taken as the S peak, and the relative retention time of peak 10 and peak 9 is calculated) and Fig.33 It shows that the relative retention time and relative peak area of ​​the three batches of cardamom kernel formula granules are within the required range.

[0191] Table 14 Relative retention time determination results of three batches of grass kernel formula granules

[0192]

[0193] Table 15 Relative peak area determination results of three batches of grass kernel formula granules

[0194]

[0195] Experimental Example 3 Establishment of the control characteristic spectrum

[0196] (1) Generation of reference feature maps

[0197] 17 batches of standard decoction of tsaoko kernel slices (lyophilized powder) (2103001Y, 2103002Y, 2103003Y, 2103004Y, 2103005Y, 2103006Y, 2103007Y, 2103008Y, 2103009Y, 2103010Y, 2103011Y, 2103013Y, 2103014Y, 2103015 Y, 230301Y, 230302Y, 230303Y) were used as test samples, and test sample solutions were prepared according to the method of Example 1; Amomum villosum control medicinal material was used to prepare Amomum villosum control medicinal material reference solution according to the method of Example 1; catechin, epicatechin, and citral reference substances were used to prepare catechin reference substance reference solution, epicatechin reference substance reference solution, and citral reference substance reference solution according to the method of Example 1.

[0198] The above solution was tested by high performance liquid chromatography of Example 1. The test results are shown in Tables 16-21 and Figure 34-35 . The measurement results show that the characteristic spectra of 17 batches of standard decoctions of Tsaoko kernel slices have 10 characteristic peaks in total, and the retention times should correspond to the 10 characteristic peaks in the chromatograms of the reference medicinal materials. The fingerprint similarity evaluation software "Chinese Herbal Chromatographic Fingerprint Similarity Evaluation System 2012 Edition" compiled by the Pharmacopoeia Committee was used to generate a control characteristic spectrum for the characteristic spectra of 17 batches of standard decoctions of Tsaoko kernel slices. The Mark peak fitting method was used for fitting the control spectrum, and the common characteristic peaks were identified. A total of 10 common characteristic peaks were identified, such as Fig.36 As shown in the control characteristic spectrum (Note: in Table 16, since the wavelength is switched to 240 nm from 30 to 45 minutes, Peak 9 is taken as the S peak, and the relative retention time of Peak 10 and Peak 9 is calculated).

[0199] Table 16 Relative retention time of reference spectrum

[0200] Peak 1 Peak 2 Peak 3 Peak 4 Peak 5(S) Peak 6 Peak 7 Peak 8 Peak 9 Peak 10 min 4.904 7.315 8.473 10.4 12.013 13.848 15.687 21.405 37.711 38.566 t / ts 0.41 0.61 0.71 0.87 1.00 1.15 1.31 1.78 1.00 1.02

[0201] Table 17 Comparison of relative peak areas of the spectrum

[0202] Peak 1 Peak 2 Peak 3 Peak 4 Peak 5(S) Peak 6 Peak 7 Peak 8 Peak 9 Peak 10 A 62.985 58.328 398.577 379.004 981.677 235.453 602.445 87.41 157.189 260.211 A / As 0.064 0.059 0.406 0.386 1.000 0.240 0.614 0.089 1.000 1.655

[0203] Table 18 Relative retention time of characteristic spectrum of Amomum villosum reference medicinal materials

[0204] Peak 1 Peak 2 Peak 3 Peak 4 Peak 5(S) Peak 6 Peak 7 Peak 8 Peak 9 Peak 10 min 4.806 7.219 8.34 10.292 11.935 13.806 15.676 21.344 37.68 38.542 t / ts 0.40 0.60 0.70 0.86 1.00 1.16 1.31 1.79 1.00 1.02

[0205] Table 19 Relative peak areas of characteristic spectra of Amomum villosum reference medicinal materials

[0206] Peak 1 Peak 2 Peak 3 Peak 4 Peak 5(S) Peak 6 Peak 7 Peak 8 Peak 9 Peak 10 A 10028 10634 48979 152348 467607 149880 382517 54502 30042 52049 A / As 0.021 0.023 0.105 0.326 1.000 0.321 0.818 0.117 1.000 1.733

[0207] Table 20 Relative retention time determination results of 17 batches of standard decoction of grass kernel slices (freeze-dried powder)

[0208]

[0209]

[0210] Table 21 Relative peak area determination results of 17 batches of standard decoction of grass kernel slices (lyophilized powder)

[0211]

[0212]

[0213] (3) Identification of characteristic peaks in the reference characteristic spectrum

[0214] Take appropriate amount of protocatechuic acid, protocatechuic aldehyde, epicatechin, proanthocyanidin B2, and citral reference substances, accurately weigh them, and add methanol to make solutions containing 30 μg per 1 ml, to prepare protocatechuic acid reference substance reference solution, protocatechuic aldehyde reference substance reference solution, epicatechin reference substance reference solution, proanthocyanidin B2 reference substance reference solution, and citral reference substance reference solution. Take appropriate amount of catechin reference substance, accurately weigh it, and add methanol to make a solution containing 20 μg per 1 ml, to prepare catechin reference substance reference solution.

[0215] The above solution was tested by high performance liquid chromatography of Example 1 to obtain a reference chromatogram, which was compared with the reference characteristic spectrum. The results are shown in Fig.37 , 10 characteristic peaks were located and identified by reference substances. The retention times of peaks 1, 2, 3, 4, and 5 corresponded to the retention times of protocatechuic acid, protocatechuic aldehyde, catechin, proanthocyanidin B2, and epicatechin reference substances, respectively. The retention time of peaks 9-10 corresponded to the retention time of citral reference substances. It was determined that peak 1 was protocatechuic acid, peak 2 was protocatechuic aldehyde, peak 3 was catechin, peak 4 was proanthocyanidin B2, peak 5 was epicatechin, and peaks 9 and 10 corresponded to citral (peaks 9 and 10 corresponded to two isomers of citral, neral and geranial).

[0216] Through the positioning and identification of reference substances, the first half is mainly composed of polyphenolic components such as protocatechuic acid, protocatechuic aldehyde, catechin, proanthocyanidin B2, epicatechin, etc., and the second half is volatile components such as citral. The same method can be used to determine the main water-soluble and volatile active ingredients in the formula granules of grass kernels. Among the water-soluble components in the front part of the characteristic spectrum, epicatechin has a large response, so epicatechin is selected as the S peak to calculate the relative retention time of each characteristic peak.

[0217] In summary, the test sample chromatogram should show 10 characteristic peaks, and should correspond to the retention time of the 10 characteristic peaks in the reference chromatogram of the reference medicinal material, among which peaks 3, 5, 9, and 10 should correspond to the retention time of the corresponding reference peaks. The peak corresponding to the epicatechin reference peak is the S peak. The relative retention time of peaks 1-4, 6-10 and S peak is calculated. The relative retention time should be within the range of ±10% of the specified value, and the specified value is: 0.41 (peak 1), 0.61 (peak 2), 0.87 (peak 4), 1.15 (peak 6), 1.31 (peak 7), 1.78 (peak 8). Peak 1 is protocatechuic acid, peak 2 is protocatechuic aldehyde, peak 3 is catechin, peak 4 is proanthocyanidin B2, peak 5 is epicatechin, and peaks 9 and 10 correspond to citral.

[0218] Experimental Example 4 Methodology Verification

[0219] 1. Precision

[0220] The same sample solution of the cardamom kernel formula granules prepared according to the method of Example 1 was taken and repeated for 6 times. The spectrum was recorded and the relative retention time and relative peak area of ​​10 characteristic peaks were measured. The results are shown in Tables 22-23. The RSDs of the relative retention times of the 10 characteristic peaks were all less than 2%; the relative peak areas of the characteristic peaks were slightly different, indicating good precision.

[0221] Table 22 Precision test relative retention time results

[0222]

[0223] Table 23 Precision test relative peak area results

[0224]

[0225]

[0226] 2. Repeatability test

[0227] The same sample of the cardamom kernel formula granules was taken for testing, and the samples were repeated 6 times according to the method of Example 1. The samples were injected and analyzed according to the chromatographic conditions recorded in Example 1, and the chromatograms were recorded. The results are shown in Tables 24-25. The RSDs of the relative retention times of the 10 characteristic peaks were all less than 2%, and the relative peak area differences of the characteristic peaks were small, indicating good repeatability.

[0228] Table 24 Relative retention time of repeatability test

[0229]

[0230] Table 25 Precision test relative peak area results

[0231]

[0232]

[0233] 3. Intermediate precision (different operators)

[0234] Three inspectors used the same equipment at different times to measure the same batch of cardamom kernel formula granules according to the test solution preparation method and chromatographic conditions described in Example 1. The relative retention time and relative peak area of ​​10 common peaks were measured and analyzed. The results are shown in Tables 26-27. The RSDs of the relative retention times of the 10 characteristic peaks were all less than 2%, indicating that the method had good intermediate precision.

[0235] Table 26 Intermediate precision relative retention time

[0236]

[0237] Table 27 Intermediate precision relative retention area

[0238]

[0239] 4. Stability test

[0240] Take the same portion of the test solution prepared in Example 1, and according to the chromatographic conditions recorded in Example 1, inject and analyze at 0, 4, 8, 12, 18, and 24 h, record the chromatogram, determine the relative retention time and relative peak area of ​​10 characteristic peaks, and analyze them. The results are shown in Tables 28-29. The relative retention time RSDs of the 10 characteristic peaks are all less than 2%, indicating that the method has good stability and can meet the needs of the determination.

[0241] Table 28 Relative retention time of stability test

[0242]

[0243] Table 29 Relative retention area of ​​stability test

[0244]

[0245] 5. Exclusivity

[0246] Preparation of negative blank solvent: Take the negative particles prepared with maltodextrin, grind them into powder, take about 0.2g, weigh accurately, put it in a stoppered conical flask, add 25ml of 70% v / v methanol aqueous solution, weigh the weight, ultrasonically treat (power 250W, frequency 40kHz) for 30 minutes, cool, make up the lost weight with 70% methanol, shake well, filter, and take the filtrate to obtain.

[0247] Accurately pipette the test solution (2 μL) and negative blank solvent (2 μL) obtained in Example 1 and inject them into the high performance liquid chromatograph respectively. Test according to the chromatographic conditions of Example 1. Fig.38 As shown in Figure 1, the results show that the negative has no interference.

[0248] 6. Durability inspection

[0249] 6.1. Investigation of different flow rates

[0250] Take the same portion of the test solution prepared in Example 1, and measure it at flow rates of 0.45 ml / min, 0.50 ml / min, and 0.55 ml / min according to the chromatographic conditions recorded in Example 1, record the chromatogram, measure the relative retention time and relative peak area at different flow rates, and analyze them. The results are shown in Tables 30-31;

[0251] Table 30 Results of relative retention time at different flow rates

[0252]

[0253] Table 31 Results of relative peak area investigation at different flow rates

[0254]

[0255] Different flow rates have a certain influence on the information content of the chromatographic peaks and the system adaptability parameters. After comparing and analyzing the above results, when the flow rates are 0.45ml / min and 0.55ml / min, the separation of Peak 2 and Peak 6 is affected. Therefore, the present invention selects a flow rate of 0.50ml / min as the best.

[0256] 6.2. Investigation of different column temperatures

[0257] Take the same portion of the test solution prepared in Example 1, and measure it according to the chromatographic conditions recorded in Example 1 at column temperatures of 25°C, 30°C, and 35°C, respectively, record the chromatogram, measure the relative retention time and relative peak area at different column temperatures, and analyze them. The results are shown in Tables 32-33;

[0258] Table 32 Results of relative retention time at different column temperatures

[0259]

[0260] Table 33 Results of relative peak area investigation at different column temperatures

[0261]

[0262] Different column temperatures have a certain influence on the information content of the chromatographic peaks and the system adaptability parameters. After comparing and analyzing the above results, when the column temperature is 25°C and 35°C, the separation of peak 2 and peak 3 is affected. Therefore, the present invention selects a column temperature of 30°C as the best.

[0263] 6.3. Investigation of different brands of chromatographic columns

[0264] The same sample solution prepared in Example 1 was taken and measured using different chromatographic columns (chromatographic column 1: Waters ACQUITY HSS T3; chromatographic column 2: Welch core-shell; chromatographic column 3: CAPCELLCORE C18) according to the chromatographic conditions recorded in Example 1. The chromatograms were recorded, and the relative retention times and relative peak areas of different chromatographic columns were measured and analyzed. The results are shown in Tables 34-35.

[0265] Table 34 Results of relative retention time investigation on different chromatographic columns

[0266]

[0267] Table 35 Results of relative peak area investigation on different chromatographic columns

[0268]

[0269] Different chromatographic columns have a certain influence on the information content of the chromatographic peak and the system adaptability parameters. After comparing and analyzing the above results, the chromatographic separation obtained by the Waters ACQUITY HSS T3 chromatographic column is good. Therefore, the present invention selects Waters ACQUITY HSS T3 (1.8 μm, 3.0×150 mm) as the best chromatographic column.

[0270] 6.4. Investigation of different chromatographs

[0271] The same sample solution prepared in Example 1 was taken and measured using different chromatographs (Chromatograph 1: Waters ACQUITY UPLC H-Class PLUS chromatographic system; Chromatograph 2: Agilent 1290 Infinity II chromatographic system; Chromatograph 3: Thermo Vanquish chromatographic system) according to the chromatographic conditions described in Example 1, and the chromatograms were recorded. The relative retention time and relative peak area of ​​different instruments were measured and analyzed. The results are shown in Tables 36-37.

[0272] Table 36 Results of relative retention time investigation on different chromatographic columns

[0273]

[0274]

[0275] Table 37 Results of relative peak area investigation on different chromatographic columns

[0276]

[0277] After comparing and analyzing the above results, the reproducibility of each chromatographic peak on different chromatographs is good, indicating that the method provided by the present invention has good durability.

[0278] Example 2

[0279] According to relevant information, the main counterfeit products of Amomum villosum are Amomum villosum, ... wild Amomum villosum, and Pseudo-Amomum villosum (Guangxi Amomum villosum), etc.

[0280] The present embodiment provides a method for distinguishing between tsaoko or tsaoko kernels or their derivative products and counterfeits, respectively using tsaoko medicinal materials, Amomum villosum Lour. (scientific name: dried mature fruit of Amomum villosum Lour., a plant of the ginger family), Amomum gagnepainii (scientific name: dried mature fruit of Amomum gagnepainii TLWu et al., a plant of the ginger family), Amomum zerumbet (scientific name: dried mature fruit of Alpinia zerumbet (Pers.) Burtt. & Smith, a plant of the ginger family), Amomum muricarpum Elm. (scientific name: dried mature fruit of Amomum muricarpum Elm., a plant of the ginger family), Alpinia katsumadai Hayata (scientific name: dried nearly mature seeds of Alpinia katsumadai Hayata, a plant of the ginger family), Amomum koenigii (scientific name: dried mature fruit of Amomum koenigii JFGmelin, a plant of the ginger family), Amomum pseudo-tsaoko (scientific name: dried mature fruit of Amomum pseudo-tsaoko, a plant of the ginger family), and Amomum tsaoko. The dried mature fruit of SQTong & Y.M.Xia) was used as the test sample, and characteristic spectra were prepared according to the method of Example 1. The characteristic spectra of each counterfeit product were compared with the characteristic spectra of Amomum villosum, and the results are shown in Figure 39-41 shown.

[0281] Compared with the control characteristic spectrum of Amomum villosum, the characteristic spectra of Amomum villosum, Amomum villosum, Amomum villosum, Amomum villosum, Amomum villosum, Amomum villosum, and Amomum villosum have basically no chromatographic peaks at the positions of peaks 7 and 8. Considering that peak 8 is smaller in the control characteristic spectrum of Amomum villosum, the relative peak area of ​​peak 8 and peak S is also smaller (0.046-0.174). Therefore, the minimum value of the relative peak area of ​​peak 7 and peak S is specified (0.390×0.8=0.312) to distinguish Amomum villosum from related counterfeits.

[0282] It can be seen that the identification method of the present invention can be used to identify cardamom or cardamom kernels or their derivatives and counterfeits. If peak 7 appears in the characteristic spectrum of the product to be identified and the relative peak area of ​​peak 7 and peak S is ≥0.31, it is cardamom or its derivatives, otherwise it is not.

[0283] Example 3

[0284] According to relevant information, the grass fruit shell should be removed when used clinically.

[0285] This embodiment provides a method for identifying tsaoko kernels or their derivatives and tsaoko in shells or their derivatives, comprising the following steps:

[0286] (1) Using the tsaoko medicinal material, tsaoko kernel slices, tsaoko kernel freeze-dried powder (standard decoction), and tsaoko (standard decoction) as test samples, respectively, prepare test sample solutions according to the method of Example 1; taking appropriate amounts of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substances, respectively, prepare 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substance solutions and meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substance solutions containing 2.5 μg per ml according to the method of Example 1.

[0287] (2) Based on Figure 42-43 As shown, the optimal absorption wavelength of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol is 192 nm, so the wavelength of 192 nm was selected for high performance liquid chromatography determination.

[0288] Take 2 μL of the test solution and the reference solution and use high performance liquid chromatography to determine the content of the sample. The chromatographic conditions are as follows: octadecylsilane bonded silica gel is used as the filler (Waters HSST3) column length 150mm, inner diameter 3.0mm, particle size 1.8μm); acetonitrile as mobile phase A, 0.1% v / v phosphoric acid solution as mobile phase B, gradient elution according to the provisions in Table 1; flow rate 0.5ml per minute; column temperature 30℃; detection wavelength 192nm. The results are shown in Tables 38-39 and Fig.44 shown.

[0289] Table 38 Comparison of calculated concentration results of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol

[0290]

[0291] Table 39 Comparison of calculated concentration results of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol

[0292]

[0293]

[0294] according to Fig.44 As shown in the figure, the detection peaks of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in the cardamom and its derivatives are relatively high; the above two components are basically not detected in the cardamom kernels and their derivatives. Therefore, meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol can be used to identify and distinguish the cardamom kernels and their derivatives from the cardamom in shells and their derivatives.

[0295] It can be seen from Tables 38-39 that the concentration of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol is more different from that of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in cardamom and its derivatives and cardamom kernels and their derivatives. Based on cost considerations, only the peak area of ​​a certain concentration of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference solution is specified to identify and differentiate the shelled cardamom.

[0296] The minimum concentration of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in samples related to tsaoko is 2.72 μg / ml, and the maximum concentration in samples related to tsaoko kernel is 1.057 μg / ml. Therefore, the concentration of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substance is determined to be 2.5 μg / ml. At a wavelength of 192 nm, the peak area of ​​the chromatographic peak of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in the sample to be tested shall not be greater than the peak area of ​​meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substance. The results are shown in Table 40.

[0297] Table 40 Comparison of peak area results of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol

[0298]

[0299]

[0300] The peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substance with a concentration of 2.5 μg / ml is 155239 μAU·S. According to Table 40, the peak areas of the Amomum villosum-related samples are 181611-377171 μAU·S, the peak areas of the Amomum villosum kernel-related samples are 10574-70568 μAU·S, and the peak areas of the Amomum villosum kernel-related derivatives are 44865-55420 μAU·S. It can be seen that the peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol reference substance with a concentration of 2.5 μg / ml can effectively distinguish the Amomum villosum kernel and its derivatives from the Amomum villosum shell.

[0301] It can be seen that the identification method of the present invention can be used to identify grass fruit kernels or their derivatives and shelled grass fruit or their derivatives. For shelled grass fruit or their derivatives, the chromatographic peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol is detected to be greater than 155239 μAU·S; for grass fruit kernels or their derivatives, the chromatographic peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol is detected to be ≤155239 μAU·S.

[0302] Obviously, the above embodiments are merely examples for the purpose of clear explanation, and are not intended to limit the implementation methods. For those skilled in the art, other different forms of changes or modifications can be made based on the above description. It is not necessary and impossible to list all the implementation methods here. The obvious changes or modifications derived from these are still within the scope of protection of the invention.

Claims

1. A method for constructing a characteristic spectrum of Amomum villosum or Amomum villosum kernel or its derivative products, characterized in that: The method includes the use of high performance liquid chromatography, and the chromatographic conditions include: Octadecylsilane bonded silica gel was used as filler, aqueous solution containing phosphoric acid was used as mobile phase B, acetonitrile was used as mobile phase A, and gradient elution was performed. The gradient elution procedure included: 0-12 minutes, the volume percentage of mobile phase A is 9% → 16%, the volume percentage of mobile phase B is 91% → 84%; 12-18 minutes, mobile phase A volume percentage is 16% → 24%, the volume percentage of mobile phase B is 84% → 76%; 18-24 minutes, the volume percentage of mobile phase A is 24% → 30%, the volume percentage of mobile phase B is 76% → 70%; 24-35 minutes, mobile phase A volume percentage is 30% → 52%, the volume percentage of mobile phase B is 70% → 48%; 35-45 minutes, the volume percentage of mobile phase A is 52% → 55%, the volume percentage of mobile phase B is 48% → 45%.

2. The construction method according to claim 1, characterized in that: The chromatographic conditions include at least one of the following: (1) The detection wavelength is 230-240 nm; optionally, 230 nm for 0 to 30 minutes and 240 nm for 30 to 45 minutes; (2) The flow rate is 0.45-0.55 ml / min; optionally, the flow rate is 0.5 ml / min; (3) Column temperature 25-35°C; optionally, column temperature 30°C; (4) The injection volume is 1-5 μL; optionally, the injection volume is 1-3 μL; further optionally, the injection volume is 2 μL; (5) The phosphoric acid concentration in the aqueous solution containing phosphoric acid in the mobile phase B is 0.05 to 0.2% v / v; optionally, the phosphoric acid concentration is 0.1% v / v; (6) The specifications of the chromatographic column are: column length 100-150 mm, inner diameter 3.0-4.6 mm, and particle size 1.8-2.7 μm; optionally, the column length is 150 mm, the inner diameter is 3.0 mm, and the particle size is 1.8 μm.

3. The construction method according to claim 1 or 2, characterized in that: The method also includes the preparation of a test sample solution, including: weighing the test sample, adding a solvent for extraction, separating the solid from the liquid, and taking a liquid, which is the test sample solution.

4. The construction method according to claim 3, characterized in that: The preparation of the test solution satisfies at least one of the following: A. The ratio of the mass of the test sample to the volume of the solvent is (0.2-1.0):25; the relationship between mass and volume is g / mL; B. The extraction method is ultrasonic extraction; C. Extraction time is ≥15 minutes, preferably 30 minutes; D. The solid-liquid separation is selected from centrifugation or membrane filtration; E. The solvent is selected from one or more of methanol, ethanol and water; preferably, it is a methanol-water solution with a volume percentage of 50-70%.

5. The construction method according to any one of claims 1 to 4, characterized in that: The construction method further comprises the steps of preparing a reference solution using at least one of catechin, epicatechin and citral, and obtaining a reference spectrum of the reference solution by detecting the reference solution using the high performance liquid chromatography method in the construction method; Preferably, each 1 mL of catechin reference solution contains 10-30 μg of catechins; preferably, 20 μg; Preferably, each 1 mL of epicatechin reference solution contains 20-40 μg of epicatechin; preferably, 30 μg; Preferably, each 1 mL of citral reference solution contains 20-40 μg of citral; preferably, 30 μg; Preferably, the solvent used in the preparation of the reference solution is selected from methanol or a methanol aqueous solution with a volume fraction of not less than 50-70%; more preferably, the solvent used in the preparation of the reference solution is selected from a 70% methanol aqueous solution; Preferably, the construction method further comprises the step of preparing a control medicinal material solution using the Amomum villosum control medicinal material, and the step of detecting the control medicinal material solution by the high performance liquid chromatography method in the construction method to obtain a control medicinal material reference atlas; Preferably, the amomum villosum or its derivative products include at least one of amomum villosum medicinal materials, amomum villosum decoction pieces or amomum villosum preparations; the amomum villosum kernels or their derivative products include at least one of amomum villosum kernel decoction pieces or amomum villosum preparations.

6. A method for determining the content of tsaoko or tsaoko kernel or its derivative products, characterized in that: include: Take the test solution and the reference solution, and respectively detect them by the high performance liquid chromatography method in the method for constructing the characteristic spectrum of the tsaoko or tsaoko kernel or its derivative products according to any one of claims 1 to 5; Wherein, the reference substance includes at least one of catechin, epicatechin and citral.

7. A method for detecting the quality of tsaoko or tsaoko kernels or their derivative products, characterized in that: The method comprises the steps of comparing the characteristic spectrum of the product to be tested with the control characteristic spectrum of the tsaoko or tsaoko kernel or a product derived therefrom; The characteristic spectrum of the product to be tested is constructed according to the construction method described in any one of claims 1 to 5; The reference characteristic spectrum of the Amomum villosum or Amomum villosum kernel or its derivative products is selected from any one of the following (1)-(4): (1) It has 10 common characteristic peaks. The peak corresponding to the reference peak of epicatechin is the S peak. The relative retention time of peaks 1-2, 4, 6-8 and S peak is calculated. The relative retention time should be within the range of ±10% of the specified value. The specified values ​​of peaks 1-2, 4 and 6-8 are: 0.41, 0.61, 0.87, 1.15, 1.31 and 1.78 respectively. (2) It has 10 common characteristic peaks, of which 4 peaks correspond to the retention times of reference peaks of catechin, epicatechin and citral respectively, and the peak corresponding to the reference peak of epicatechin is the S peak. The relative retention times of peaks 1-2, peak 4 and peaks 6-8 and the S peak are calculated, and the relative retention times should be within the range of ±10% of the specified values. The specified values ​​of peaks 1-2, peak 4 and peak 6-8 are: 0.41, 0.61, 0.87, 1.15, 1.31, 1.78 respectively; optionally, peak 1 corresponds to protocatechuic acid; peak 2 corresponds to protocatechuic aldehyde; peak 3 corresponds to catechin; peak 4 corresponds to proanthocyanidin B2; peak 5 (S) corresponds to epicatechin; peaks 9 and 10 correspond to citral; (3) Characteristic spectrum of the Amomum villosum and / or Amomum villosum kernels and / or Amomum villosum derivatives obtained by the construction method according to any one of claims 1 to 5 using a single batch or multiple batches of Amomum villosum and / or Amomum villosum kernels and / or Amomum villosum derivatives as test samples; (4) Using multiple batches of cardamom and / or cardamom kernels and / or their derivatives as test samples, the characteristic spectrum obtained according to the construction method described in any one of claims 1 to 5 is used to prepare a control characteristic spectrum by using the average value or median method.

8. A method for distinguishing between tsaoko or tsaoko kernels or their derivatives and counterfeits, characterized in that: include: The test is carried out according to the quality testing method of the tsaoko or tsaoko kernel or its derivative products as described in claim 7; wherein, for the tsaoko or tsaoko kernel or its derivative products, the relative peak area of ​​peak 7 and peak S should be within the specified range, and the specified value is: not less than 0.31; Optionally, the counterfeit products include one or more of Amomum villosum or its derivatives, Alpinia cyrtonema or its derivatives, Amomum villosum or its derivatives, Amomum villosum or its derivatives; preferably, the Amomum villosum includes one or more of Amomum villosum yangchunensis, Amomum villosum longifolia, Amomum villosum niuginosa, and Amomum villosum fragrant.

9. A method for distinguishing between tsaoko kernels or their derivatives and tsaoko in shell or their derivatives, characterized in that: include: The step of detecting the characteristic spectrum of the product to be tested by high performance liquid chromatography in the construction method according to any one of claims 1 to 5; The chromatographic conditions of the high performance liquid chromatography method include a detection wavelength of 192-194 nm; The reference solution also includes a reference solution containing 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol; For the tsaoko kernel or its derivative products, the peak area of ​​the chromatographic peak of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in the characteristic spectrum is ≤ the peak area of ​​the chromatographic peak of the corresponding reference solution of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol; For shelled amomum villosum or its derivative products, the chromatographic peak area of ​​1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol in the characteristic spectrum is greater than the chromatographic peak area of ​​the corresponding reference solution of 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol and / or meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol.

10. The method for identifying the tsaoko kernels or their derivatives and the tsaoko in shells or their derivatives according to claim 9, characterized in that: Each 1 mL of the reference solution contains 1.6-2.5 μg of meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol or 1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol; Preferably, for shelled cardamom or its derivative products, the chromatographic peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol is detected to be greater than 155239 μAU·S; for cardamom kernel or its derivative products, the chromatographic peak area of ​​the meso-1,7-bis-(4-hydroxyphenyl)-3,5-heptanediol is detected to be ≤155239 μAU·S.

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