Kelp (kelp) formula granule preparation characteristic spectrum construction method, kelp (kelp) formula granule preparation quality detection method and counterfeit product identification method

Abstract

The invention relates to the field of traditional Chinese medicine detection and quality detection, in particular to a kelp (kelp) formula granule preparation characteristic spectrum construction method, a quality detection method and a counterfeit product identification method. The method for constructing the characteristic chromatogram of the kelp preparation comprises the step of detecting by adopting ultra-high performance liquid chromatography under the chromatographic conditions that octadecylsilane chemically bonded silica is used as a filling agent, acetonitrile is used as a mobile phase A, a 0.03-0.07% phosphoric acid aqueous solution is used as a mobile phase B, and gradient elution is carried out; according to the specific chromatogram construction method, the quality of the kelp preparation can be comprehensively controlled, the constructed specific chromatogram has more characteristic peaks and identified components, the characteristic peak information of the sample can be comprehensively reflected, the peak pattern is good, the purity is high, the separation degree is good, the precision is high, and the reproducibility is good; the method can be used for identifying the kelp preparation and four common adulterants thereof.

Description

Technical Field

[0001] The present invention relates to the field of traditional Chinese medicine detection and quality inspection, and in particular to a method for constructing a characteristic spectrum of a kelp (seaweed) formula granule preparation, a quality inspection method and a method for identifying counterfeit and mixed products. Background Art

[0002] Konbu is the dried thallus of Laminaria japonica Aresch. of the Laminariaceae family or Ecklonia kurome Okam. of the Pterophyceae family. The original plant is a perennial algae. Konbu has a long history of medicinal use in my country and is a commonly used medicinal material. This product is cold in nature and salty in taste. It enters the liver, stomach, and kidney meridians. It belongs to the category of Chinese medicine for softening and dispersing nodules. Its effects are to eliminate phlegm, soften and dissipate nodules, and promote diuresis and reduce swelling. It is used for goiter, scrofula, testicular swelling and pain, and edema caused by phlegm and fluid retention.

[0003] Kelp has a wide range of clinical applications. For example, it can be used in combination with Fritillaria, Citrus aurantium, and Pinellia tuber to treat the initial stage of goiter, which may be hard or swollen. It can be used in combination with Citrus aurantium, Aloe vera, and Sea Powder to clear the liver, relieve depression, and resolve phlegm. It can be used in combination with Ginseng, Angelica sinensis, and Rehmannia glutinosa to treat goiter that has been around for a long time and has weak Qi and blood. It can be used in combination with Notopterygium incisum, Saposhnikovia divaricata, Seaweed, and Forsythia suspensa to treat aversion to cold and fever, and the initial stage of scrofula. It can be used in combination with Ginseng, Angelica sinensis, and Cyperus rotundus to treat scrofula caused by stagnation of liver Qi and insufficient Qi and blood. It can be used in combination with Bupleurum, Seaweed, Gentiana scabra, and Tripterygium wilfordii to treat scrofula that is all over the jaw, hard and not ulcerated, and with excessive heat and toxicity. It can be used in combination with Seaweed, Citrus aurantium, Cinnamomum cassia, and Corydalis yanhusuo to treat testicular pain caused by cold and dampness in the lower Jiao, Qi stagnation, and blood stasis. It can be used in combination with Tangerine peel, Pinellia tuber, and Poria cocos to treat plum pit qi. It can be used to treat beriberi edema when combined with Radix Angelicae Dahuricae, Radix Stephaniae Tetrandrae, Semen Plantaginis, etc. It is used in modern clinics to treat retinal concussion, thyroid disease, hypertension, chronic pelvic inflammatory disease, cancer, breast hyperplasia, phlebitis, viral non-icteric hepatitis, constipation, vitreous opacity, senile cataracts, etc. Modern pharmacodynamics results show that this product has multiple pharmacological effects such as affecting the cardiovascular system, lowering blood lipids, lowering blood sugar, anticoagulation, anti-radiation, anti-tumor, antitussive, antiasthmatic, anti-mutation, anti-oxidation, antiviral and antibacterial, and affecting immune function.

[0004] Common counterfeits of kelp include the dried thallus of Ulva actuca L, Ulva pertusakjellm from the Ulvaceae family, the dried thallus of Undaria Pinnatifida (Harv.) Sur from the Pterophyceae family, the dried thallus of Brasenia schreberi JFGmel. from the Brasenaceae family, and the dried algae of Sargassum pallidum (Turn.) C.Ag (commonly known as "big-leaf seaweed") or Sargassum fusiforme (Harv.) Setch. (commonly known as "small-leaf seaweed") from the Sargassumaceae family.

[0005] At present, the characteristic spectrum detection method in the existing kelp formula granule standard stipulates fewer characteristic peaks, fewer identified components of kelp, and the identified components are nucleosides, which are common components of most medicinal materials, which is not conducive to the quality control of preparations such as kelp formula granules, and it is also impossible to distinguish between mixed and counterfeit products. Compared with kelp medicinal materials, kelp formula granules and other preparations have undergone water decoction and extraction and other process steps, and their active ingredients are very different from those of kelp medicinal materials. The characteristic spectrum detection method of kelp medicinal materials is not applicable to preparations such as kelp formula granules, and the important characteristic peaks and index components of preparations such as kelp formula granules are difficult to separate. Summary of the invention

[0006] Therefore, the technical problem to be solved by the present invention is to overcome the problem that the characteristic spectrum of kelp preparations in the prior art has fewer characteristic peaks and fewer identifying components, making it difficult to comprehensively control its quality and unable to mix counterfeits. To this end, the present invention provides a method for constructing a characteristic spectrum of kelp preparations, a quality detection method and a method for identifying counterfeits. The constructed characteristic spectrum has more characteristic peaks, more identifying components, a high peak type, and high purity, and can be distinguished from 4 types of counterfeits.

[0007] To this end, the present invention provides the following technical solutions:

[0008] The invention provides a method for constructing a characteristic spectrum of a kelp preparation, comprising detection by ultra-high performance liquid chromatography, wherein the chromatographic conditions include: using octadecylsilane bonded silica gel as a filler, using acetonitrile as a mobile phase A, and using a 0.03-0.07% phosphoric acid aqueous solution as a mobile phase B, and performing gradient elution, wherein the elution procedure is as follows: 0-10 minutes, mobile phase A is 15 v / v%→27 v / v%, and mobile phase B is 85 v / v%→73 v / v%; 10-25 minutes, mobile phase A is 27 v / v%→87 v / v% v%, mobile phase B is 73v / v%→13v / v%; 25-28 minutes, mobile phase A is 87v / v%, mobile phase B is 13v / v%; 28-29 minutes, mobile phase A is 87v / v%→100v / v%, mobile phase B is 13v / v%→0v / v%; 29-40 minutes, mobile phase A is 100v / v%, mobile phase B is 0v / v%; 40-45 minutes, mobile phase A is 100v / v%→15v / v%, mobile phase B is 0v / v%→85v / v%.

[0009] In some embodiments, the chromatographic conditions further include at least one of the following conditions:

[0010] 1) The flow rate is 0.9-1.1 ml per minute; optionally, the flow rate is 1.0 ml per minute;

[0011] 2) The column temperature is 29-31°C; optionally, the column temperature is 30°C;

[0012] 3) The detection wavelength is 210nm to 330nm; optionally, the detection wavelength is 210nm at 0 to 13min and 28 to 45min, and 330nm at 13 to 28min;

[0013] 4) The injection volume is 15 μl; Optionally, the injection volume is 15 μl;

[0014] 5) The specifications of the chromatographic column are: column length 250 mm, inner diameter 4.6 mm, particle size 5 μm; Optionally, the specifications of the chromatographic column are: column length 250 mm, inner diameter 4.6 mm, particle size 5 μm;

[0015] 6) Mobile phase B is 0.05% phosphoric acid aqueous solution.

[0016] In some embodiments, the preparation of the test solution includes: taking the test sample, adding an extraction solvent, extracting, separating the solid from the liquid, drying, and adding a re-dissolving solvent to dissolve;

[0017] And / or, the test sample includes water extract, standard decoction or formula granules of Kelp.

[0018] In some embodiments, the preparation of the test solution includes at least one of the following conditions:

[0019] 1) The extraction solvent is water, 30-100 v / v% methanol or its aqueous solution, or 30-100 v / v% ethanol or its aqueous solution; optionally, methanol is selected;

[0020] 2) The extraction method used in the extraction step is ultrasonic extraction or reflux extraction;

[0021] 3) The extraction time in the extraction step is 30-50min;

[0022] 4) The mass volume ratio of the test sample and the extraction solvent is 0.5-2g:20ml;

[0023] 5) The re-dissolving solvent is 50-100 v / v% methanol or its aqueous solution; optionally, 70 v / v% methanol aqueous solution is selected;

[0024] 6) The mass volume ratio of the test sample and the reconstitution solvent is 0.5~2g:2ml.

[0025] In some embodiments, the preparation of a reference solution is further included, comprising:

[0026] Preparation of control medicinal material solution: taking control medicinal material, adding extraction solvent, extracting, solid-liquid separation, drying, adding re-dissolving solvent to dissolve;

[0027] And / or, preparation of reference solution: take linoleic acid reference, add solvent to make 1 ml of solution containing 0.05-0.15 mg; optionally, 1 ml of solution containing 0.1 mg; optionally, the solvent is 50-100 v / v% methanol or its aqueous solution; optionally, 70 v / v% methanol aqueous solution is selected.

[0028] In some embodiments, the preparation of the control medicinal material solution includes at least one of the following conditions:

[0029] 1) The extraction solvent is water, 30-100 v / v% methanol or its aqueous solution, or 30-100 v / v% ethanol or its aqueous solution;

[0030] 2) The extraction method used in the extraction step is ultrasonic extraction or reflux extraction;

[0031] 3) The number of extractions in the extraction step is 1 to 2 times;

[0032] 3) The extraction time in the extraction step is 30-50min;

[0033] 4) The mass volume ratio of the reference medicinal material and the extraction solvent is 0.5-2 g: 20-50 ml;

[0034] 5) The re-dissolving solvent is 50-100 v / v% methanol or its aqueous solution; optionally, 70 v / v% methanol aqueous solution is selected;

[0035] 6) The mass volume ratio of the control medicinal material and the reconstitution solvent is 0.5-2g:2ml.

[0036] The present invention provides a method for detecting the content of a kelp preparation, wherein a test solution and a reference solution are taken and respectively detected by ultra-high performance liquid chromatography in the method for constructing a characteristic spectrum of the kelp preparation;

[0037] Among them, the reference substance includes linoleic acid.

[0038] The present invention provides a quality detection method for a kelp preparation, comprising the steps of comparing a characteristic spectrum of a sample to be tested with a control characteristic spectrum of a kelp preparation;

[0039] The characteristic spectrum of the sample to be tested is constructed according to the construction method;

[0040] The control characteristic spectrum of the kelp preparation is selected from any one of the following (1)-(2):

[0041] (1) It has 10 common characteristic peaks, and peak 10 corresponding to the peak of the linoleic acid reference substance is the S peak. The relative retention time of peaks 1-9 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 1-9 are: 0.27, 0.29, 0.34, 0.67, 0.88, 0.92, 0.94, 0.95, 0.98;

[0042] (2) Using at least one batch of kelp preparation standard product and obtaining the characteristic spectrum according to the construction method, a control characteristic spectrum is prepared by using the average value or median method.

[0043] In some embodiments, in (1), peak 2 corresponds to isoarachidonic acid; peak 3 corresponds to isoarachidonic acid; peak 6 corresponds to arachidonic acid; peak 7 corresponds to α-linolenic acid; peak 8 corresponds to γ-linolenic acid; peak 9 corresponds to arachidonic acid; and peak 10 (S) corresponds to linoleic acid.

[0044] The present invention provides a method for distinguishing a kelp preparation and its counterfeit products, comprising: performing detection according to the quality detection method of the kelp preparation;

[0045] Compared with the characteristic spectrum of the control kelp preparation or the characteristic spectrum of the kelp preparation standard as the test product, when the adulterated product is wakame or its derivatives, its characteristic spectrum contains at least one chromatographic peak at 35 minutes to 40 minutes and lacks chromatographic peak 1;

[0046] Compared with the characteristic spectrum of the control kelp preparation or the characteristic spectrum of the kelp preparation standard as the test product, when the adulterated product is microphylla or its derivatives, its characteristic spectrum lacks chromatographic peak 8 and lacks chromatographic peak 1;

[0047] Compared with the control characteristic spectrum of kelp preparation or the characteristic spectrum of kelp preparation standard as the test product, when the adulterated product is large-leaf seaweed or its derivatives, the peak areas of peaks 2 and 3 in its characteristic spectrum are respectively greater than the peak areas of chromatographic peaks 4 to 10, and the peak area of ​​peak 4 is greater than the peak area of ​​peak 6, and chromatographic peak 1 is missing;

[0048] Compared with the control characteristic spectrum of kelp preparation or the characteristic spectrum of kelp preparation standard as the test product, when the adulterated product is Brasenia schrenkiana or its derivatives, the peak area of ​​peak 7 in its characteristic spectrum is greater than the peak area of ​​peak 6, and the chromatographic peak 1 is missing.

[0049] In the present invention, the kelp preparation includes water extract, standard decoction or formula granules of kelp.

[0050] The technical solution of the present invention has the following advantages:

[0051] 1. A method for constructing a characteristic spectrum of a kelp preparation provided by the present invention comprises detecting by ultra-high performance liquid chromatography, wherein the chromatographic conditions comprise: using octadecylsilane bonded silica gel as a filler, acetonitrile as a mobile phase A, and 0.03-0.07 v / v% phosphoric acid aqueous solution as a mobile phase B, and performing gradient elution, wherein the elution procedure is as follows: 0-10 minutes, mobile phase A is 15 v / v% → 27 v / v%, mobile phase B is 85 v / v% → 73 v / v%; 10-25 minutes, mobile phase A is 27 v / v% → 87 v / v%, mobile phase B is 73 v / v% → 13 v / v%; 25-28 minutes, mobile phase A is 87 v / v%, mobile phase B is 10 v / v% → 15 v / v%; Mobile phase B is 13v / v%; 28-29 minutes, mobile phase A is 87v / v%→100v / v%, mobile phase B is 13v / v%→0v / v%; 29-40 minutes, mobile phase A is 100v / v%, mobile phase B is 0v / v%; 40-45 minutes, mobile phase A is 100v / v%→15v / v%, mobile phase B is 0v / v%→85v / v%; The above-mentioned characteristic spectrum construction method can comprehensively control the quality of kelp preparations, and the constructed characteristic spectrum has more characteristic peaks and identification components, which can comprehensively reflect the characteristic peak information of the sample, and has good peak shape, high purity, good separation, high accuracy and good reproducibility;

[0052] Furthermore, the above method can be used to identify kelp preparations and their four common counterfeits, and can enhance the specific identification of kelp preparations;

[0053] Furthermore, the above method can also determine the content of kelp preparations, and determine the index components such as isothiocyanate, dithiocyanate, arachidonic acid, α-linolenic acid, γ-linolenic acid, arachidonic acid, and linoleic acid. BRIEF DESCRIPTION OF THE DRAWINGS

[0054] 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.

[0055] Figure 1 is the characteristic spectrum of the test sample in Example 1 of the present invention; (1) is the characteristic spectrum of the test sample; (2) is the characteristic spectrum of the reference sample; (3) is the characteristic spectrum of the reference medicinal material;

[0056] Figure 2 This is a comparative characteristic spectrum of the kelp formula granules in Example 2 of the present invention;

[0057] Figure 3 is the delay test result in Example 3 of the present invention;

[0058] Figure 4 is the characteristic spectrum under different extraction solvent conditions in Example 4 of the present invention;

[0059] Figure 5 is a characteristic spectrum under different extraction methods in Example 4 of the present invention;

[0060] Figure 6 It is the characteristic spectrum at different extraction times in Example 4 of the present invention;

[0061] Figure 7 It is the characteristic spectrum under different extraction dosages in Example 4 of the present invention;

[0062] Figure 8 is the characteristic spectrum under different re-dissolution solvents in Example 4 of the present invention;

[0063] Fig. 9 is the characteristic spectrum of the test solution in Example 5 of the present invention;

[0064] Fig.10 is the characteristic spectrum of the linoleic acid reference substance solution in Example 5 of the present invention;

[0065] Fig.11 It is the characteristic spectrum of the auxiliary material negative solution in Example 5 of the present invention;

[0066] Fig.12 is the characteristic spectrum of the blank solvent in Example 5 of the present invention;

[0067] Fig.13 is the characteristic spectrum of each mixed and counterfeit product in Example 6 of the present invention;

[0068] Fig.14 yes Fig.13 A partial enlarged view of

[0069] Fig.15 It is the characteristic spectrum of the comparative example of the present invention. DETAILED DESCRIPTION

[0070] 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.

[0071] 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.

[0072] Instruments, reagents and test drugs

[0073] Instruments: Shimadzu LC-2010A HT high performance liquid chromatograph; ME104E electronic balance (Mettler Toledo Scientific Instruments (Shanghai) Co., Ltd.), JY20002 electronic balance (Shanghai Sunny Hengping Instrument Co., Ltd.), BSA124S electronic balance (Sartorius Scientific Instruments (Beijing) Co., Ltd.), XP26 electronic balance (Mettler Toledo Scientific Instruments (Shanghai) Co., Ltd.), KQ-500DB ultrasonic cleaner (Kunshan Ultrasonic Instrument Co., Ltd.).

[0074] Chromatographic column:

[0075] Agilent ZORBAX Eclipse XDB-C18 (4.6mm*250mm, 5μm);

[0076] Agilent ZORBAX Extend-C18 (4.6mm*250mm, 5μm);

[0077] Agilent ZORBAX Eclipse Plus C18 (4.6mm*250mm, 5μm).

[0078] Drug testing:

[0079] Kelp formula granules (batch numbers: K402CP01, K402CP02, K402CP03, self-made by Beijing Kangrentang Pharmaceutical Co., Ltd.); the preparation method of Kelp formula granules is as follows: take Kelp, heat and reflux extraction twice, add 14 times the weight of water of Kelp for the first time, heat and reflux extraction for 1 hour, filter, add 12 times the weight of water of Kelp for the second time, heat and reflux extraction for 1 hour, filter, combine the filtrate, concentrate the filtrate to a relative density of 1.0 g / mL (measured at 60°C), spray dry, add auxiliary material maltodextrin to the dry powder obtained by spray drying (the total weight of the dry powder after adding maltodextrin accounts for 45.4% of the amount of Kelp fed), mix well, and then dry granulate to make granules.

[0080] Standard decoction of kelp (batch number: K402BJ01-15, self-made by Beijing Kangrentang Pharmaceutical Co., Ltd.) Preparation method of standard decoction of kelp: take 100-150g of kelp slices, put them in a decoction pot, add 16 times the amount of water to the slices for the first decoction, soak in cold water for 30 minutes, boil over high heat (500W), boil over low heat (200W) for 30 minutes, filter with a 150-mesh filter cloth while hot, cool quickly and set aside; add 14 times the amount of water to the slices for the second decoction, boil over high heat (500W), boil over low heat (200W) for 20 minutes, filter with a 150-mesh filter cloth while hot, cool quickly and set aside; combine the filtrate, concentrate under reduced pressure (temperature set at 65°C, vacuum degree -0.1MPa) to a solid-liquid ratio of about 1:1 (relative density of 1.05-1.10 (65°C)), freeze-dry and make into fine powder.

[0081] Kelp (Laminaria japonica) control medicinal material (Batch number: AY300005-202305, Foshan Ao Yu Biotechnology Co., Ltd.).

[0082] Linoleic acid reference substance (batch number: 111622-202105, purity 99.6%, China Food and Drug Inspection Institute);

[0083] Reagents: acetonitrile (Merk) and phosphoric acid (Fisher Scientific) were of chromatographic grade; water was distilled water (Watsons); methanol and ethyl acetate were of analytical grade.

[0084] Example 1 Method for constructing characteristic spectrum of kelp preparation

[0085] This embodiment provides a method for constructing a characteristic spectrum of a kelp preparation, including using ultra-high performance liquid chromatography to detect:

[0086] The chromatographic conditions are as follows: octadecylsilane bonded silica gel as filler (column length 250 mm, inner diameter 4.6 mm, particle size 5 μm, chromatographic column: ZORBAX Extend-C18); acetonitrile as mobile phase A, 0.05 v / v% phosphoric acid as mobile phase B, gradient elution as specified in the following table; flow rate 1.0 ml per minute; column temperature 30°C; detection wavelengths 210 nm (0-13 min and 28-45 min) and 330 nm (13-28 min). The theoretical plate number calculated based on the linoleic acid peak should be no less than 5000.

[0087] Table 1. Gradient elution program

[0088] Time (minutes) Mobile phase A (v / v%) Mobile phase B (v / v%) 0～10 15→27 85→73 10～25 27→87 73→13 25～28 87 13 28～29 87→100 13→0 29～40 100 0 40～45 100→15 0→85

[0089] Preparation of reference solution: Take about 2.0g of kelp reference medicinal material, add 50ml of water, reflux extraction for 30 minutes, cool, filter, and concentrate the filtrate to dryness under reduced pressure. Add 20ml of methanol to the residue, ultrasonically treat (power 250W, frequency 40kHz) for 40 minutes, take out, cool, filter, evaporate the filtrate to dryness, and dissolve it in 2ml of 70% methanol solution as the reference medicinal material solution. Take an appropriate amount of linoleic acid reference substance, accurately weigh it, and add 70% methanol to make a solution containing 0.1mg per 1ml as the linoleic acid reference substance solution.

[0090] Preparation of test solution: Take an appropriate amount of kelp formula granules (batch numbers K402CP01, K402CP02, K402CP03), grind them into powder, take about 1.0 g, weigh it, put it in a stoppered conical flask, add 20 ml of methanol, ultrasonically treat it (power 250 W, frequency 40 kHz) for 40 minutes, cool it, filter it, evaporate the filtrate to dryness, and dissolve it in 2 ml of 70% methanol solution to obtain the solution.

[0091] Determination method: Accurately pipette 15 μl of reference solution and test solution respectively, inject into liquid chromatograph, and determine.

[0092] The characteristic spectrum of the measurement is as follows

[0093] Table 2. Relative retention time of three batches of kelp formula particles

[0094]

[0095] The characteristic spectrum of the test sample, the characteristic spectrum of the reference sample and the characteristic spectrum of the reference medicinal material are as follows: Figure 1 As shown in the figure and the table above, there are 10 characteristic peaks in the characteristic spectrum of the test sample, and the retention time of the 10 characteristic peaks in the characteristic spectrum of the control corresponds to that of the peak 10, of which peak 10 should correspond to the retention time of the linoleic acid reference peak, and the peak corresponding to the linoleic acid reference peak is the S peak. The relative retention time of the other characteristic peaks and the S peak is calculated, and the relative retention time should be within the range of ±10% of the specified value, and the specified value is: 0.27 (peak 1), 0.29 (peak 2), 0.34 (peak 3), 0.67 (peak 4), 0.88 (peak 5), 0.92 (peak 6), 0.94 (peak 7), 0.95 (peak 8), 0.98 (peak 9). Among them, peak 2 corresponds to iso-schizophylline; peak 3 corresponds to schizophylline; peak 6 corresponds to arachidonic acid; peak 7 corresponds to α-linolenic acid; peak 8 corresponds to γ-linolenic acid; peak 9 corresponds to arachidonic acid; peak 10 (S) corresponds to linoleic acid. According to the above method, the test sample can be quality tested. The test sample presents 10 characteristic peaks, which is relatively large. Seven components can be identified, so its quality can be fully controlled. The characteristic spectrum has good separation, stable baseline, good peak shape and high purity.

[0096] Example 2 Establishment of the comparative characteristic spectrum of kelp preparation

[0097] 15 batches of kelp standard decoction and 3 batches of kelp formula granules were taken respectively, and their characteristic spectra were obtained according to the method of Example 1. The characteristic spectra of 15 batches of kelp standard decoction are shown in the table below, and the characteristic spectra of 3 batches of kelp formula granules are shown in Table 2 in Example 1. The "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System" (2021.1 version) was used, and one of the characteristic spectra was used as a reference spectrum. The median was calculated to obtain the control spectrum of the kelp preparation as shown in Figure 2 As shown, common peaks were identified, and 10 common peaks were identified.

[0098] Table 3. Retention time of 15 batches of kelp standard decoction

[0099]

[0100]

[0101] Table 4. Peak areas and relative peak areas of 15 batches of kelp standard decoctions

[0102]

[0103]

[0104] Table 5. Relative retention time of characteristic spectra of kelp formula particles

[0105]

[0106] Table 6. Relative peak areas of characteristic spectra of kelp formula particles

[0107]

[0108]

[0109] The characteristic peaks in the reference characteristic spectrum of the kelp formula granules established above were identified. After positioning and identifying each peak using the reference substance, peak 2 was identified as iso-kalpol; peak 3 was identified as kalpol; peak 6 was identified as arachidonic acid; peak 7 was identified as α-linolenic acid; peak 8 was identified as γ-linolenic acid; peak 9 was identified as arachidonic acid; peak 10 was identified as linoleic acid, and peak 10 (linoleic acid) was determined as the S peak.

[0110] Finally, it was determined that 10 characteristic peaks should be present in the chromatogram of the test sample, among which peak 10 should correspond to the retention time of the linoleic acid reference substance peak. The peak corresponding to the linoleic acid reference substance is the S peak. The relative retention time of peak 1, peak 2, peak 3, peak 4, peak 5, peak 6, peak 7, peak 8, peak 9 and S peak was calculated, and the relative retention time should be within ±10% of the specified value. The specified values ​​are: 0.27 (peak 1), 0.29 (peak 2), 0.34 (peak 3), 0.67 (peak 4), 0.88 (peak 5), 0.92 (peak 6), 0.94 (peak 7), 0.95 (peak 8), and 0.98 (peak 9). Among them, peak 2 corresponds to iso-schizonepeta lactone; peak 3 corresponds to iso-schizonepeta lactone; peak 6 corresponds to arachidonic acid; peak 7 corresponds to α-linolenic acid; peak 8 corresponds to γ-linolenic acid; peak 9 corresponds to arachidonic acid; peak 10 (S) corresponds to linoleic acid

[0111] Example 3 Delay Test

[0112] Take kelp formula granules (batch number K402CP01) according to the test solution preparation method in Example 1, and then detect according to the chromatographic conditions in Example 1, and record 2 times the mobile phase elution time. The results are as follows Figure 3 shown.

[0113] The results showed that no obvious hysteresis peak appeared after 40 minutes, and this chromatographic method met the analysis requirements.

[0114] Example 4 System suitability test

[0115] Kelp formula granules (batch number K402CP01) were prepared according to the test solution preparation method in Example 1, and then the system suitability test was performed according to the chromatographic conditions in Example 1, with the linoleic acid peak as a reference. The results are shown in the following table, and it can be concluded that the theoretical plate number, tailing factor and retention time of the linoleic acid peak all meet the analysis requirements.

[0116] Table 7. System suitability test results

[0117]

[0118]

[0119] Example 5 Investigation of preparation of test solution

[0120] 1. Investigation of extraction solvent

[0121] Weigh 9 portions of kelp formula granules (batch number: K402CP01), 1.0 g each, grind, weigh accurately, put in a stoppered conical flask, accurately add water, 30% methanol, 50% methanol, 70% methanol, methanol, 30% ethanol, 50% ethanol, 70% ethanol, 20 ml of ethanol, weigh, ultrasonically treat (power 250W, frequency 40kHz) for 40 minutes, cool, add corresponding solvent to make up, filter and evaporate to dryness, dissolve with 2 ml of corresponding solvent, and obtain. Accurately draw 15 μl of the filtrate, inject into a high performance liquid chromatograph, and measure the peak area of ​​each peak according to the chromatographic conditions in Example 1. The results are as shown in the following table and Figure 4 The chromatogram and data analysis showed that when the extraction solvent was ethanol, the proposed characteristic peak substances were difficult to extract, and the extraction efficiency of 70% methanol was basically the same as that of methanol; however, during the sample preparation process, 70% methanol would extract more sugars, making it difficult to redissolve. Taking all factors into consideration, in order to ensure sufficient extraction and easy preservation of organic solvents, methanol was selected as the best extraction solvent for the preparation of the characteristic spectrum of kelp formula particles.

[0122] Table 8. Extraction solvent inspection table

[0123] Extraction solvent Peak 1 Peak 2 Peak 3 Peak 4 Peak 5 Peak 6 Peak 7 Peak 8 Peak 9 Peak 10 water 88377 379303 415718 31264 267992 185131 74741 28525 109218 38338 30% methanol 130971 347168 384818 56910 231334 126115 51219 39483 57588 21887 50% methanol 172329 377798 410489 71138 604503 268308 101759 88886 69650 29280 70% methanol 185201 430447 470335 85173 1402586 1092852 408035 357485 508198 211611 Methanol 369853 354603 369824 61984 1481711 1147561 434799 376496 550329 229555 30% ethanol 41836 374057 403183 67989 409698 237505 93131 77205 112516 41336 50% ethanol 65950 303631 347516 63562 1003248 785694 294176 256590 373303 154939 70% ethanol 48046 344587 361547 59633 1000630 814864 300038 267222 391182 158225 Ethanol 8234 60623 53584 34194 712237 628242 234182 193395 301232 124825

[0124] 2. Investigation of extraction methods

[0125] Weigh 2 portions of kelp (seaweed) formula granules (batch number: K402CP01), 1.0 g each, grind, weigh accurately, place in a stoppered conical bottle, accurately add 20 ml of methanol, weigh, ultrasonically treat (power 250 W, frequency 40 kHz) for 40 minutes and heat under reflux for 40 minutes, cool, add methanol to make up the weight, filter and evaporate to dryness, dissolve with 2 ml of methanol, and obtain. Accurately draw 15 μl of the filtrate, inject into a high performance liquid chromatograph, and measure the peak area of ​​each peak according to the chromatographic conditions in Example 1. The results are as shown in the following table and Figure 5 The chromatogram and data analysis showed that there was no significant difference between the reflux extraction characteristic spectrum and ultrasonic extraction, and the ultrasonic peak area was higher. After comprehensive consideration, ultrasonic extraction was selected as the best extraction method for the preparation of the characteristic spectrum of kelp formula particles.

[0126] Table 9. Investigation of extraction methods

[0127] Extraction method Peak 1 Peak 2 Peak 3 Peak 4 Peak 5 Peak 6 Peak 7 Peak 8 Peak 9 Peak 10 Ultrasound 369853 354603 369824 61984 1481711 1147561 434799 376496 550329 229555 Reflow 392721 296866 310853 59575 1162598 927555 357750 311379 476600 225988

[0128] 3. Extraction time investigation

[0129] Weigh three portions of kelp (seaweed) formula granules (batch number: K402CP01), about 1.0 g each, grind finely, weigh accurately, place in a stoppered conical bottle, accurately add 20 ml of methanol, weigh the weight, ultrasonically treat (power 250 W, frequency 40 kHz) for 30, 40, and 50 minutes, cool, add methanol to make up the weight, filter and evaporate to dryness, and dissolve with 2 ml of methanol to obtain. Accurately draw 15 μl of the filtrate, inject into a high performance liquid chromatograph, and measure the peak area of ​​each peak according to the chromatographic conditions in Example 1. The results are as shown in the following table and Figure 6 The results show that the peak areas of the characteristic peaks in the characteristic spectra obtained at different extraction times are basically the same. In order to ensure that the peak area of ​​each chromatographic peak is moderate, the extraction time of 40 minutes is determined as the optimal extraction time.

[0130] Table 10. Extraction time inspection table

[0131] Extraction time Peak 1 Peak 2 Peak 3 Peak 4 Peak 5 Peak 6 Peak 7 Peak 8 Peak 9 Peak 10 30min 247897 214197 218573 40860 1016914 793508 298610 258001 398547 156355 40min 369853 354603 369824 61984 1481711 1147561 434799 376496 550329 229555 50min 298122 254104 263015 46260 1081251 865212 318527 273269 421612 168243

[0132] 4. Investigation of extraction dosage

[0133] Weigh three portions of kelp (seaweed) formula granules (batch number: K402CP01), about 0.5, 1.0, and 2.0 g, respectively, grind, weigh accurately, place in a stoppered conical bottle, accurately add 20 ml of methanol, weigh, ultrasonically treat (power 250 W, frequency 40 kHz) for 40 minutes, cool, add methanol to make up the weight, filter and evaporate to dryness, and dissolve with 2 ml of methanol to obtain. Accurately draw 15 μl of the filtrate, inject into a high performance liquid chromatograph, and measure according to the chromatographic conditions in Example 1 to measure the peak area of ​​each peak. The results are as shown in the following table and Figure 7 The results showed that the total peak areas of the samples with sampling amounts of 0.5g, 1g and 2g were compared and found that there was no significant difference after the sampling amount conversion, indicating that the extraction efficiency of the proposed characteristic peaks in the samples with sampling amounts of 0.5g, 1g and 2g was basically the same, indicating that the above-mentioned extraction parameters can stably extract the characteristic peak substances of 1g. Taking comprehensive considerations, 1g was determined to be the optimal sampling amount for the characteristic spectrum of kelp formula particles.

[0134] Table 11. Extraction dosage survey table

[0135]

[0136]

[0137] 5. Investigation of reconstitution solvent

[0138] Weigh three portions of kelp (seaweed) formula granules (batch number: K402CP01), about 1.0 g each, grind finely, weigh accurately, place in a stoppered conical flask, accurately add 20 ml of methanol, weigh the weight, ultrasonically treat (power 250 W, frequency 40 kHz) for 40 minutes, cool, add methanol to make up the weight, filter and evaporate to dryness, dissolve with 2 ml of methanol, 70% methanol, and 50% methanol respectively, and obtain. Accurately draw 15 μl of the filtrate, inject into a high performance liquid chromatograph, and measure the peak area of ​​each peak according to the chromatographic conditions in Example 1. The results are as shown in the following table and Figure 8 During the experiment, it was found that the sample dissolved in methanol would crystallize, and 50% methanol would affect the peak area of ​​the characteristic peak. After comprehensive consideration, 70% methanol was determined to be the best dissolution solvent for the characteristic spectrum of kelp formula particles.

[0139] Table 12, Reconstitution Solvent Investigation Table

[0140] Reconstitution solvent Peak 1 Peak 2 Peak 3 Peak 4 Peak 5 Peak 6 Peak 7 Peak 8 Peak 9 Peak 10 Methanol 369853 354603 369824 61984 1481711 1147561 434799 376496 550329 229555 70% methanol 380065 374555 410694 91333 1400743 1068036 413598 357904 478894 214283 50% methanol 405308 409026 428405 84290 687955 341327 131709 114635 110369 44810

[0141] Example 5 Methodology Verification

[0142] 1. Exclusivity

[0143] Take linoleic acid reference substance (prepared according to the linoleic acid reference substance solution in Example 1), test solution (kelp formula granules (batch number: K402CP01), prepared according to the test solution in Example 1), blank solvent (70v / v% methanol), and auxiliary material negative sample, and measure according to the chromatographic conditions in Example 1. The results are as follows: Figures 9 to 12 As shown, it is illustrated that the method of the present invention has strong specificity.

[0144] Preparation of auxiliary material negative solution: Take an appropriate amount of maltodextrin, grind it into powder, take about 1.0g, weigh it, put it in a stoppered conical flask, add 20ml of methanol, ultrasonically treat it (power 250W, frequency 40kHz) for 40 minutes, cool it, filter it, evaporate the filtrate to dryness, and dissolve it in 2ml of 70% methanol solution to obtain it.

[0145] 2. Precision

[0146] Weigh 6 portions of kelp (seaweed) formula granules (batch number: K402CP01), grind them, and measure them according to the chromatographic conditions in Example 1 to obtain their characteristic spectrum. Take peak No. 10 as the reference peak to calculate its relative peak area and relative retention time. And calculate RSD. The results are shown in the following table. The relative peak area RSD of each characteristic peak is in the range of 0.77-3.91%, and the relative peak area difference is slightly large. Considering that the error of redissolution in the sample preparation method is large, it may cause the area difference of each characteristic peak to be large, so the relative peak area is not specified. The relative retention time RSD of each characteristic peak is in the range of 0-0.11%, and the relative retention time difference is small, indicating that the repeatability of the characteristic spectrum is good.

[0147] Table 13. Retention time and relative retention time table of repeatability test of kelp formula granules

[0148]

[0149] Table 14. Peak area and relative peak area of ​​repeatability test of kelp

[0150]

[0151] 2. Intermediate precision

[0152] Using a high performance liquid chromatograph and a PDA detector, 6 portions of kelp formula granules (batch number: K402CP01) were taken, and the chromatographic conditions in Example 1 were measured to obtain its characteristic spectrum. Peak No. 10 was used as the reference peak to calculate its relative peak area and relative retention time. And RSD was calculated. The results are shown in the following table. In the characteristic spectrum obtained by using a high performance liquid chromatograph (PDA detector), the relative retention time RSD of each characteristic peak was 0.02-0.05%, and the relative peak area RSD was in the range of 1.20-2.03%. The relative retention time RSD between different instruments (two high performance liquid chromatographs (PDA detectors)) was 0-0.20%, and the relative peak area was slightly different between different instruments, with an RSD of 0.95-18.20%, which may be related to different detectors, so the relative peak area is not specified. The experimental results show that the relative retention time between different instruments of the characteristic spectrum method meets the analysis requirements.

[0153] Table 15. Retention time and relative retention time table of intermediate precision study of kelp

[0154]

[0155] Table 16. Peak area and relative peak area of ​​kelp (seaweed) intermediate precision test

[0156]

[0157]

[0158] 3. Stability

[0159] Take kelp formula granules (batch number: K402CP01), grind them into powder, prepare the test solution according to Example 1, and measure them according to the chromatographic conditions of Example 1 at 0, 2, 4, 8, 12, and 24 hours respectively to obtain its characteristic spectrum. Take peak No. 10 as the reference peak, calculate its relative peak area and relative retention time. And calculate RSD. The results are shown in the following table. The results of the stability experiment show that the RSD of the relative peak area is in the range of 0.34-1.25%, and the RSD of the relative retention time of each characteristic peak is in the range of 0-0.32%, indicating that the relative retention time of the characteristic spectrum method meets the analysis requirements when the stability is investigated.

[0160] Table 17. Retention time and relative retention time table of stability study of kelp

[0161]

[0162] Table 18. Peak area and relative peak area of ​​stability test of kelp

[0163]

[0164]

[0165] 4. Durability

[0166] 4.1 Investigation of different column temperatures:

[0167] Take kelp formula granules (batch number: K402CP01) and prepare the test solution according to Example 1, and measure them at different column temperatures (29°C, 30°C and 31°C) according to the chromatographic conditions of Example 1 to investigate the durability of the experimental method to the column temperature. The results are as shown in the following table. In the chromatograms obtained at different column temperatures, the relative retention time RSD of each characteristic peak is in the range of 0.03 to 0.66%, and the relative peak area RSD is in the range of 0.56 to 2.88%. The results show that the column temperature has little effect on the relative peak area and relative retention time. The present invention does not specify the relative peak area, so the method has good durability to the column temperature.

[0168] Table 19. Retention time and relative retention time table of kelp at different column temperatures

[0169]

[0170] Table 20. Peak area and relative peak area of ​​kelp at different column temperatures

[0171]

[0172] 4.2 Investigation of different flow rates:

[0173] Take kelp formula granules (batch number: K402CP01), prepare the test solution according to Example 1, and use different flow rates (0.9ml / min, 1.0ml / min and 1.1ml / min) to measure according to the chromatographic conditions in Example 1 to investigate the durability of the experimental method for flow rate. The results are shown in the following table. As can be seen from the results, the relative retention time RSD of each characteristic peak in the chromatogram obtained at different flow rates is in the range of 0.10-5.38% (in line with the specified value ±10%), and the relative peak area RSD is in the range of 0.04-1.76%. This method has good durability for column temperature.

[0174] Table 21. Retention time and relative retention time table of kelp at different flow rates

[0175]

[0176] Table 22. Peak area and relative peak area of ​​kelp at different flow rates

[0177]

[0178] 4.3 Investigation of different chromatographic columns:

[0179] Take kelp formula granules (batch number: K401CP01), prepare the test solution according to Example 1, and use different types of chromatographic columns (column 1: Agilent ZORBAX Eclipse XDB-C18 (4.6mm*250mm, 5μm), column 2: Agilent ZORBAX Extend-C18 (4.6mm*250mm, 5μm), column 3: Agilent ZORBAX Eclipse PlusC18 (4.6mm*250mm, 5μm)) to measure according to the chromatographic conditions of Example 1, and investigate the durability of the experimental method for different types of chromatographic columns. The results are shown in the following table. The results show that the chromatograms obtained under different chromatographic columns have a relative retention time RSD of each characteristic peak in the range of 0.18 to 7.88%, and the relative peak area RSD is larger, with an RSD range of 0.27 to 3.64%. It shows that this method has good durability for different chromatographic columns.

[0180] Table 23. Retention time and relative retention time table of kelp on different chromatographic columns

[0181]

[0182] Table 24. Peak area and relative peak area of ​​kelp in different chromatographic columns

[0183]

[0184] Example 6 Method for Identifying Kelp Preparations and Counterfeit Products

[0185] This embodiment provides a method for distinguishing kelp preparations from counterfeit and adulterated products. The sample to be tested is:

[0186] Large leaf seaweed (dried algae of Sargassum pallidum (Turn.) C.Ag of the Sargassum family) sample: Take an appropriate amount of large leaf seaweed, grind it into powder, take about 1.0 g, weigh it, put it in a stoppered conical flask, add 20 ml of methanol, ultrasonically treat it (power 250 W, frequency 40 kHz) for 40 minutes, cool it, filter it, evaporate the filtrate to dryness, and dissolve it in 2 ml of 70v / v% methanol solution to obtain the product.

[0187] Small-leaf seaweed (dried algae of Sargassum fusiforme (Harv.) Setch.) sample: Take an appropriate amount of small-leaf seaweed, grind it into powder, take about 1.0 g, weigh it, put it in a stoppered conical flask, add 20 ml of methanol, ultrasonically treat it (power 250 W, frequency 40 kHz) for 40 minutes, cool it, filter it, evaporate the filtrate to dryness, and dissolve it in 2 ml of 70v / v% methanol solution to obtain the product.

[0188] Undaria pinnatifida (Harv.) Sur) sample: Take an appropriate amount of Undaria pinnatifida, grind it into powder, take about 1.0 g, weigh it, put it in a stoppered conical flask, add 20 ml of methanol, ultrasonically treat it (power 250 W, frequency 40 kHz) for 40 minutes, cool it, filter it, evaporate the filtrate to dryness, and dissolve it in 2 ml of 70v / v% methanol solution to obtain the product.

[0189] Water shield (dried thallus of Brasenia schreberi JFGmel. of the Brasenaceae family) sample: Take an appropriate amount of water shield, grind it into fine powder, take about 1.0 g, weigh it, put it in a stoppered conical flask, add 20 ml of methanol, ultrasonically treat it (power 250 W, frequency 40 kHz) for 40 minutes, cool it, filter it, evaporate the filtrate to dryness, and dissolve it in 2 ml of 70v / v% methanol solution to obtain the product.

[0190] Take the above-mentioned samples to be tested (each sample is 3 batches) and construct characteristic spectra according to the method in Example 1. The characteristic spectra of one batch of the exemplary samples are as follows: Fig.13 The comparison results of the characteristic spectrum of the kelp formula granules are as follows: Fig.14 As shown. You can see:

[0191] Compared with the control characteristic spectrum of kelp preparation, when the counterfeit product is kelp or its derivatives, its characteristic spectrum contains at least one chromatographic peak at 35 minutes to 40 minutes, and lacks chromatographic peak 1; the comparison results of the remaining batches of samples are consistent with the above.

[0192] Compared with the control characteristic spectrum of kelp preparation, when the adulterated product is small-leaf seaweed or its derivatives, its characteristic spectrum lacks chromatographic peak 8 and chromatographic peak 1; the comparison results of the remaining batches of samples are consistent with the above.

[0193] Compared with the control characteristic spectrum of kelp preparations, when the counterfeit product is large-leaf seaweed or its derivatives, the peak areas of peaks 2 and 3 in its characteristic spectrum are respectively greater than the peak areas of chromatographic peaks 4 to 10, and the peak area of ​​peak 4 is greater than the peak area of ​​peak 6, and chromatographic peak 1 is missing; the comparison results of the remaining batches of samples are consistent with the above.

[0194] Compared with the control characteristic spectrum of kelp preparation, when the adulterated product is Brasenia schrenkiana or its derivatives, the peak area of ​​peak 7 in its characteristic spectrum is greater than the peak area of ​​peak 6, and the chromatographic peak 1 is missing. The comparison results of the remaining batches of samples are consistent with the above.

[0195] Therefore, the present invention can distinguish the kelp preparation from adulterated products.

[0196] Comparative Example

[0197] Kelp (kelp) formula granules (batch number: K401CP01) were tested according to the test sample preparation method and chromatographic conditions in "Research on UPLC Fingerprint and Chemical Pattern Recognition of Kelp" (Huang Huahua, Zhang Yiping, Lv Shishi, Journal of Chinese Medicinal Materials, Vol. 43, No. 7, July 2020). The results are as follows Fig.15 As shown, the characteristic peaks are basically lost.

[0198] 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 therefrom are still within the scope of protection of the invention.

Claims

1. A method for constructing a characteristic spectrum of a kelp preparation, characterized in that: The method includes ultra-high performance liquid chromatography detection, and the chromatographic conditions include: Octadecylsilane bonded silica gel was used as filler, acetonitrile was used as mobile phase A, and 0.03-0.07 v / v% phosphoric acid aqueous solution was used as mobile phase B. Gradient elution was performed. The elution procedure was as follows: 。 2. The method for constructing a characteristic spectrum of a kelp preparation according to claim 1, wherein: The chromatographic conditions also include at least one of the following conditions: 1) The flow rate is 0.9-1.1 ml per minute; optionally, the flow rate is 1.0 ml per minute; 2) The column temperature is 29-31°C; optionally, the column temperature is 30°C; 3) The detection wavelength is 210nm to 330nm; optionally, the detection wavelength is 210nm at 0 to 13min and 28 to 45min, and 330nm at 13 to 28min; 4) The injection volume is 15 μl; Optionally, the injection volume is 15 μl; 5) The specifications of the chromatographic column are: column length 250 mm, inner diameter 4.6 mm, particle size 5 μm; Optionally, the specifications of the chromatographic column are: column length 250 mm, inner diameter 4.6 mm, particle size 5 μm; 6) Mobile phase B is 0.05% phosphoric acid aqueous solution.

3. The method for constructing a characteristic spectrum of a kelp preparation according to claim 1 or 2, characterized in that: It also includes the preparation of a test solution, including: taking the test sample, adding an extraction solvent, extracting, separating the solid from the liquid, drying, and adding a re-dissolving solvent to dissolve; And / or, the test sample includes water extract, standard decoction or formula granules of Kelp.

4. The method for constructing a characteristic spectrum of a kelp preparation according to claim 3, characterized in that: The preparation of the test solution includes at least one of the following conditions: 1) The extraction solvent is water, 30-100 v / v% methanol or its aqueous solution, or 30-100 v / v% ethanol or its aqueous solution; optionally, methanol is selected; 2) The extraction method used in the extraction step is ultrasonic extraction or reflux extraction; 3) The extraction time in the extraction step is 30-50min; 4) The mass volume ratio of the test sample and the extraction solvent is 0.5-2g:20ml; 5) The re-dissolving solvent is 50-100 v / v% methanol or its aqueous solution; optionally, 70 v / v% methanol aqueous solution is selected; 6) The mass volume ratio of the test sample and the reconstitution solvent is 0.5~2g:2ml.

5. The method for constructing a characteristic spectrum of a kelp preparation according to any one of claims 1 to 4, characterized in that: Also included is the preparation of reference solutions, including: Preparation of control medicinal material solution: taking control medicinal material, adding extraction solvent, extracting, solid-liquid separation, drying, adding re-dissolving solvent to dissolve; And / or, preparation of reference solution: take linoleic acid reference, add solvent to make 1 ml of solution containing 0.05-0.15 mg; optionally, 1 ml of solution containing 0.1 mg; optionally, the solvent is 50-100 v / v% methanol or its aqueous solution; optionally, 70 v / v% methanol aqueous solution is selected.

6. The method for constructing a characteristic spectrum of a kelp preparation according to claim 5, characterized in that: The preparation of the control medicinal material solution includes at least one of the following conditions: 1) The extraction solvent is water, 30-100 v / v% methanol or its aqueous solution, or 30-100 v / v% ethanol or its aqueous solution; 2) The extraction method used in the extraction step is ultrasonic extraction or reflux extraction; 3) The number of extractions in the extraction step is 1 to 2 times; 3) The extraction time in the extraction step is 30-50min; 4) The mass volume ratio of the reference medicinal material and the extraction solvent is 0.5-2 g: 20-50 ml; 5) The re-dissolving solvent is 50-100 v / v% methanol or its aqueous solution; optionally, 70 v / v% methanol aqueous solution is selected; 6) The mass volume ratio of the control medicinal material and the reconstitution solvent is 0.5-2g:2ml.

7. A method for detecting the content of kelp preparation, characterized in that: Taking the test solution and the reference solution, respectively, and detecting them by the ultra-high performance liquid chromatography method in the method for constructing the characteristic spectrum of the kelp preparation according to any one of claims 1 to 6; Among them, the reference substance includes linoleic acid.

8. A quality detection method for kelp preparation, characterized in that: The method comprises the steps of comparing the characteristic spectrum of the sample to be tested with the control characteristic spectrum of the kelp preparation; The characteristic spectrum of the sample to be tested is constructed according to the construction method described in any one of claims 1 to 6; The control characteristic spectrum of the kelp preparation is selected from any one of the following (1)-(2): (1) It has 10 common characteristic peaks, and peak 10 corresponding to the peak of the linoleic acid reference substance is the S peak. The relative retention time of peaks 1-9 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 1-9 are: 0.27, 0.29, 0.34, 0.67, 0.88, 0.92, 0.94, 0.95, 0.98; (2) Using at least one batch of kelp preparation standard product and obtaining the characteristic spectrum according to the construction method described in any one of claims 1 to 6, a control characteristic spectrum is prepared by the average value or median method.

9. The quality detection method of kelp preparation according to claim 8, characterized in that: In (1), peak 2 corresponds to iso-schizotyle; peak 3 corresponds to iso-schizotyle; peak 6 corresponds to arachidonic acid; peak 7 corresponds to α-linolenic acid; peak 8 corresponds to γ-linolenic acid; peak 9 corresponds to arachidonic acid; and peak 10 (S) corresponds to linoleic acid.

10. A method for distinguishing a kelp preparation from its counterfeits, characterized in that: The method comprises testing the kelp preparation according to claim 8 or 9; Compared with the characteristic spectrum of the control kelp preparation or the characteristic spectrum of the kelp preparation standard as the test product, when the adulterated product is wakame or its derivatives, its characteristic spectrum contains at least one chromatographic peak at 35 minutes to 40 minutes and lacks chromatographic peak 1; Compared with the characteristic spectrum of the control kelp preparation or the characteristic spectrum of the kelp preparation standard as the test product, when the adulterated product is microphylla or its derivatives, its characteristic spectrum lacks chromatographic peak 8 and lacks chromatographic peak 1; Compared with the control characteristic spectrum of kelp preparation or the characteristic spectrum of kelp preparation standard as the test product, when the adulterated product is large-leaf seaweed or its derivatives, the peak areas of peaks 2 and 3 in its characteristic spectrum are respectively greater than the peak areas of chromatographic peaks 4 to 10, and the peak area of ​​peak 4 is greater than the peak area of ​​peak 6, and chromatographic peak 1 is missing; Compared with the control characteristic spectrum of kelp preparation or the characteristic spectrum of kelp preparation standard as the test sample, when the adulterated product is water shield or its derivatives, the peak area of ​​peak 7 in its characteristic spectrum is greater than the peak area of ​​peak 6, and the chromatographic peak 1 is missing.

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