A detection method for crane lice and its quality control method

Abstract

The present invention provides a method for detecting and quality controlling lice. The method comprises obtaining a characteristic spectrum using high-performance liquid chromatography (HPLC). The HPLC conditions are as follows: a chromatographic column using octadecylsilane bonded silica gel as a filler; methanol as mobile phase A; and 0.2% aqueous phosphoric acid as mobile phase B, eluting according to a specific gradient elution procedure. The present method effectively overcomes the problems of prior art detection methods, such as a small number of characteristic peaks, an unstable baseline, and low resolution. The method provides chromatographic conditions with a greater number of characteristic peaks, a more stable baseline, and improved resolution, effectively providing the prerequisites for quality control of lice-producing granules.

Description

Technical Field

[0001] The present invention relates to the field of traditional Chinese medicine detection, and in particular to a detection method for louse and a quality control method thereof. Background Art

[0002] According to literature reports, the main chemical components of schisandra chinensis are sesquiterpenes (such as tianmingjing lactone and tianmingjing lactone alcohol), fatty acids, and organic acids (such as chlorogenic acid). Since schisandra chinensis granules are made from substances extracted after boiling, the water solubility of the chemical components should be considered, and organic acids should be included as the main chemical components in the characteristic spectrum of the standard decoction of schisandra chinensis slices and the intermediates of the granules.

[0003] However, the characteristic spectrum of the lice formula granules disclosed in the prior art does not use organic acid compounds as a reference, and its characteristic peaks are relatively few, the separation between each characteristic peak is not high, and the baseline is uneven, which makes it unable to be effectively used for the quality control of the lice formula granules. Summary of the Invention

[0004] Therefore, the technical problem to be solved by the present invention is that the detection method disclosed in the prior art has the problems of small number of characteristic peaks, low separation of characteristic peaks, and uneven baseline; thus, a detection method for louse and its quality control method are provided to solve the above problems.

[0005] In order to solve the above problems, the technical solutions provided by the present invention are as follows:

[0006] A method for detecting louse of cranes, comprising using high performance liquid chromatography to obtain a characteristic spectrum, wherein the chromatographic conditions of the high performance liquid chromatography are:

[0007] Chromatographic column: Octadecylsilane bonded silica gel as filler; methanol as mobile phase A, 0.2% phosphoric acid aqueous solution as mobile phase B, elution according to the following gradient elution program:

[0008]

[0009] Among them, a is 0 to 5, b is 90 to 95, c is 95 to 105, and cb≥5.

[0010] Among the chromatographic conditions, the flow rate is 0.9-1.1 ml / min; and / or the column temperature is 28-32° C.; and / or the detection wavelength is 240 nm; and / or the theoretical plate number calculated based on the chlorogenic acid peak should be no less than 5000.

[0011] The present invention also includes the following steps of preparing the test solution: taking the test substance, accurately weighing it, adding a solvent, treating it, cooling it, adding a solvent to make up for the lost weight, shaking it evenly, filtering it, and taking a filtrate to obtain the solution.

[0012] The treatment method is heating reflux or ultrasound;

[0013] And / or, the solvent is methanol, ethanol, water or methanol-water solution.

[0014] The ultrasonic treatment time is 30-60 min; and / or the ultrasonic power is 250-300 W and the frequency is 40 kHz;

[0015] And / or, the heating reflux treatment time is 30-60 minutes.

[0016] The added amount of the solvent is 50-250 ml / g.

[0017] The injection volume of the test solution is 5-20 μl.

[0018] The chromatographic column is Agilent pursuit XRs 5 C18, Tnature C18 or YMC C18; the column length is 250 mm, the inner diameter is 4.6 mm, and the particle size is 5 μm.

[0019] A quality control method for crane lice based on the above detection method.

[0020] The quality control method includes: the characteristic spectrum of the test sample obtained by the high performance liquid chromatography method includes at least 8 characteristic peaks; among them, peak 2 is chlorogenic acid; peak 2 is the S peak, and the relative retention time of each of the other characteristic peaks should be within ±10% of the specified value; the specified values ​​of the relative retention times of the 8 characteristic peaks are 0.35, 1.00, 1.04, 1.08, 1.27, 1.59, 1.66, and 2.05, respectively.

[0021] The technical solution of the present invention has the following advantages:

[0022] The present invention provides a method for detecting crane lice. By optimizing chromatographic conditions, especially the coordination of the gradient elution program and the mobile phase, more characteristic peaks can be effectively detected, and the separation between the characteristic peaks is good, and the overall baseline is stable, thereby effectively providing the prerequisite for the quality control of crane lice. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the specific embodiments or the description of the prior art. Obviously, the drawings described below are some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0024] Figure 1 This is the characteristic spectrum of the Heshi formula granules in Example 1 of the present invention;

[0025] Figure 2 This is a chromatogram of the test sample of the Heshi formula granules in Example 2 of the present invention;

[0026] Figure 3 1 is a chromatogram of a negative sample of the Heshi formula granules in Example 2 of the present invention;

[0027] Figure 4 is a chromatogram of gradient elution procedure 1 in Example 3 of the present invention;

[0028] Figure 5 is a chromatogram of gradient elution procedure 2 in Example 3 of the present invention;

[0029] Figure 6 is a chromatogram of gradient elution procedure 3 in Example 3 of the present invention;

[0030] Figure 7 is a chromatogram of gradient elution procedure 4 in Example 3 of the present invention;

[0031] Figure 8 is a chromatogram of gradient elution procedure 5 in Example 3 of the present invention;

[0032] Figure 9 is the chromatogram of Agilent pursuit XRs 5 C18 in Example 3 of the present invention;

[0033] Figure 10 is the chromatogram of Tnature C18 in Example 3 of the present invention;

[0034] Figure 11 is the chromatogram of YMC C18 in Example 3 of the present invention;

[0035] Figure 12 This is the chromatogram at a column temperature of 20° C. in Example 3 of the present invention;

[0036] Figure 13 This is the chromatogram at a column temperature of 25° C. in Example 3 of the present invention;

[0037] Figure 14 This is a chromatogram at a column temperature of 30° C. in Example 3 of the present invention. DETAILED DESCRIPTION

[0038] If no specific experimental steps or conditions are specified in the examples, the conventional experimental steps or conditions described in the literature in this field can be used. If the manufacturer of the reagents or instruments is not specified, they are all commercially available conventional reagents.

[0039] 1. Instruments and equipment:

[0040] Chromatograph: Shimadzu high performance liquid chromatography, including LC-20AD binary pump, SIL-20A autosampler, SPD-20A detector, chromatography workstation;

[0041] Mettler Toledo NewClassic MS 100,000th balance, Jing Tian FA2044A 10,000th balance;

[0042] SB-5200DT ultrasonic cleaner (Ningbo Xinzhi).

[0043] 2. Reagents

[0044] Methanol, acetonitrile, and phosphoric acid were of chromatographic grade, and water was ultrapure water; other reagents were of analytical grade.

[0045] 3. Drug testing

[0046] Chlorogenic acid reference substance: purchased from China Food and Drug Inspection Institute, batch number 110753-202119, purity: 96.3%;

[0047] Control medicinal materials of lice: purchased from China Food and Drug Inspection Institute, batch number: 121131-202102

[0048] Crane lice formula granules: batch number: 2209001Y, 2209002Y, 2209003Y.

[0049] Example 1

[0050] A method for detecting crane lice can effectively obtain a characteristic spectrum of crane lice. The specific acquisition process is as follows:

[0051] 1. Chromatographic conditions and system suitability

[0052] The chromatographic conditions and system suitability experiments were as follows: octadecylsilane bonded silica gel was used as the filler (Tnature C18 column, column length 250 mm, inner diameter 4.6 mm, particle size 5 μm); methanol was used as mobile phase A, 0.2% aqueous solution was used as mobile phase B, and gradient elution was performed according to the provisions in Table 1 below; the flow rate was 1.0 ml per minute; the column temperature was 30°C; the detection wavelength was 240 nm, and the number of theoretical plates calculated based on the chlorogenic acid peak should be no less than 5000.

[0053] Table 1 Gradient elution table

[0054]

[0055] Preparation of reference solution: Take 0.8 g of the control medicinal material of the Chinese gentian, add 50 ml of water, heat and reflux for 60 minutes, filter, evaporate the filtrate to dryness, add 25 ml of 50% methanol to the residue, sonicate (power 250 W, frequency 40 kHz) for 30 minutes, cool, shake well, filter, and use the filtrate as the reference medicinal material solution. Accurately weigh an appropriate amount of chlorogenic acid reference substance and add 50% methanol to prepare a reference solution containing 70 μg per 1 ml. This is used as the reference solution.

[0056] Preparation of test solution: Take an appropriate amount of the product, grind it into powder, take about 0.1 g, place it in a stoppered conical flask, add 25 ml of 50% methanol, ultrasonically treat (power 250 W, frequency 40 kHz) for 30 minutes, cool, shake well, filter, and take the filtrate.

[0057] Determination method: Accurately pipette 10 μl of reference solution and test solution respectively, inject into liquid chromatograph, and determine.

[0058] The characteristic spectrum of the Heli formula granules obtained under the above chromatographic conditions of the present invention is as follows: Figure 1 The characteristic spectrum of the Heshi formula granules should show eight characteristic peaks, and their retention times should correspond to the eight characteristic peaks in the chromatogram of the reference medicinal material. Peak 2 should correspond to the retention time of the chlorogenic acid reference peak. The relative retention times of each characteristic peak and the S peak should be calculated. The relative retention times should be within ±10% of the specified values: 0.35 (peak 1), 1.04 (peak 3), 1.08 (peak 4), 1.27 (peak 5), 1.59 (peak 6), 1.66 (peak 7), and 2.05 (peak 8).

[0059] The specific process is as follows:

[0060] In this embodiment, 18 batches of standard decoction of lice slices (lyophilized powder) (batch numbers: 2103001Y, 2103002Y, 2103003Y, 2103004Y, 2103005Y, 2103006Y, 2103007Y, 2103008Y, 2103009Y, 2103010Y, 2103011Y, 2103012Y, 2103013Y, 2103014Y , 2103015Y, 211101Y, 211102Y, 211103Y) and three batches of standard decoctions (lyophilized powder) thereof were used as test solutions. The relative retention time and relative peak area of ​​each characteristic peak were obtained through 21 batches of test solutions. The test results are shown in Tables 2 and 3 below.

[0061] Table 2

[0062]

[0063]

[0064]

[0065] Table 3

[0066]

[0067]

[0068] The fingerprint test results of 18 batches of standard decoctions of Heshi medicinal pieces and 3 batches of Heshi formula granules were analyzed. The fingerprint similarity evaluation software "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System 2012 Edition" compiled by the Pharmacopoeia Committee was used to generate a control characteristic spectrum, and the 8 chromatographic peaks with good separation of the characteristic spectrum of Heshi formula granules were confirmed as follows Figure 1 shown.

[0069] The reference spectrum obtained in this example shows eight characteristic peaks, which should correspond to the retention times of the eight characteristic peaks in the chromatogram of the reference medicinal material. Peak 2 corresponds to the retention time of the chlorogenic acid reference peak. The chlorogenic acid reference peak is used as the S peak, and the relative retention time of each characteristic peak and the S peak is within ±10% of the specified value; the specified value is: 0.35 (peak 1), 1.04 (peak 3), 1.08 (peak 4), 1.27 (peak 5), 1.59 (peak 6), 1.66 (peak 7), and 2.05 (peak 8).

[0070] In order to verify the accuracy of the characteristic spectrum, three batches of Heshi formula granules were taken and operated according to the same measurement method as in Example 1 to obtain the relative peak area and relative retention time of each characteristic peak. The specific measurement results are shown in Tables 4 and 5 below.

[0071] Table 4

[0072]

[0073] Table 5

[0074]

[0075]

[0076] The above test results show that the relative retention times of the characteristic peaks of the three batches of Heshi formula granules are all within the specified value range.

[0077] Example 2

[0078] 1. System applicability

[0079] The test solution and the reference solution were prepared according to the test solution preparation method, and HPLC analysis was performed according to the chromatographic conditions described in Example 1 to verify whether the chromatographic conditions and system adaptability of the standard decoction of Helix spp. were applicable to the Helix spp. formula granules, and whether the negative sample of Helix spp. formula granules (prepared with maltodextrin according to the test solution preparation method) would cause interference. The specific test results are as follows: Figure 2-Figure 3 and as shown in Table 6 below.

[0080] Table 6

[0081]

[0082] The above test results show that the chromatographic method system has good adaptability for the test samples and has no interference with negative samples, and can be used as a detection method for the characteristic spectrum of the granules of the lice formula.

[0083] 2. Validation of analytical methods

[0084] 2.1 Precision

[0085] Take the same test solution of the Heshi formula granules and repeat the injection 6 times according to the chromatographic conditions in Example 1. Record the chromatogram, measure the relative retention time and relative peak area of ​​8 characteristic peaks, and analyze them. The results are shown in the table below.

[0086] Table 7 Precision relative retention time test results

[0087]

[0088]

[0089] Table 8 Precision relative peak area test results

[0090]

[0091] From the results in the above table, we can see that the RSDs of the relative retention time and relative peak area of ​​the characteristic peaks are small, indicating good precision.

[0092] 2.3 Method repeatability test

[0093] The same sample of the Heshi formula granules was taken and replicated 6 times. The sample was injected and analyzed according to the chromatographic conditions in Example 1. The chromatogram was recorded, and the relative retention time and relative peak area of ​​8 characteristic peaks were measured and analyzed. The results are shown in the table below. The RSDs of the relative retention time and relative peak area of ​​the characteristic peaks were small, indicating good repeatability.

[0094] Table 9 Repeatability relative retention time test results

[0095]

[0096]

[0097] Table 10 Repeatability relative peak area test results

[0098]

[0099] 2.3 Intermediate precision (different operators)

[0100] Two inspectors, at different times, used the same equipment to measure the same batch of Heshi formula granules according to the chromatographic conditions in Example 1. The relative retention times and relative peak areas of the eight common peaks were measured and analyzed. The results are shown in the table below.

[0101] Table 11 Intermediate precision relative retention time test results

[0102]

[0103]

[0104] Table 12 Intermediate precision relative peak area test results

[0105]

[0106] As can be seen from the above table, the RSDs of relative retention time and relative peak area are both small, indicating that the intermediate precision of this method is good.

[0107] 2.4 Stability investigation

[0108] Take the same test solution of the Heshi formula granules and inject the sample for analysis at 0, 4, 8, 12, 18, and 24 hours according to the chromatographic conditions in Example 1. Record the chromatogram, measure the relative retention time and relative peak area of ​​the 8 characteristic peaks, and analyze them. The results are shown in the table below.

[0109] Table 13 Stability relative retention time test results

[0110]

[0111]

[0112] Table 14 Stability relative peak area test results

[0113]

[0114] It can be seen from the above table that the RSDs of the relative retention time and relative peak area of ​​each characteristic peak are small, indicating that the method has good stability and can meet the needs of determination.

[0115] Example 3

[0116] 1. Detection results of different gradient elution programs

[0117] The gradient elution programs in Tables 15 to 19 below were used for detection, and the test results were as follows: Figure 4-Figure 8 shown.

[0118] Table 15 Gradient elution program 1

[0119]

[0120] Table 16 Gradient elution program 2

[0121]

[0122]

[0123] Table 17 Gradient elution program 3

[0124]

[0125] Table 18 Gradient elution program 4

[0126]

[0127] Table 19 Gradient elution program 5

[0128]

[0129] The above results show that: when gradient elution program 4 is used, the separation between characteristic peaks 3 and 4 is relatively poor, while when gradient elution programs 1-3 and 5 are used, the separation of characteristic peaks in the obtained chromatogram is relatively good; among them, gradient elution program 5 has better overall separation of characteristic peaks, better peak shape, and relatively shorter detection time. Therefore, gradient elution program 5 is preferred in the present invention.

[0130] 2. Test results of different chromatographic columns

[0131] The same batch of test solution was taken and analyzed according to the determination method in Example 1 using Agilent pursuit XRs5 C18 (250mm×4.6mm, 5μm), Tnature C18 (250mm×4.6mm, 5μm), and YMC C18 (250mm×4.6mm, 5μm) chromatographic columns, respectively. The chromatograms were recorded and the separation effect of each characteristic peak was analyzed. The chromatograms are shown in Figure 1. Figures 9-11 shown.

[0132] pass Figures 9-11The results show that the separation and retention time of chromatographic peaks are different for different chromatographic columns, and Tnature C18 has a better effect. Therefore, the fixed chromatographic column of the present invention is: Tnature C18 (250 mm×4.6 mm, 5 μm).

[0133] 3. Investigation of different mobile phase concentrations

[0134] The same test solution was taken and analyzed at 0.18%, 0.20%, and 0.22% phosphoric acid concentrations according to the determination method in Example 1. The relative retention time and relative peak area of ​​each characteristic peak and peak S were examined. The results are shown in the following table.

[0135] Table 20

[0136]

[0137] Table 21

[0138]

[0139] From the results in the above table, it can be seen that when the phosphoric acid concentration changes, the separation effect of peaks 6 and 7 is greatly affected. This method requires a fixed phosphoric acid concentration of 0.2% as the characteristic spectrum of the helix formula particles to determine the phosphoric acid concentration in the mobile phase.

[0140] 4. Investigation of different column temperatures

[0141] Take the same sample solution and analyze it at different column temperatures of 20℃, 25℃ and 28-32℃ according to the determination method in Example 1 to obtain chromatograms. The chromatograms at 20℃, 25℃ and 30℃ are as follows: Figure 12-14 As shown, the relative retention time and relative peak area of ​​each characteristic peak and peak S in the chromatogram between 28-32℃ are shown in the following table.

[0142] Table 22

[0143]

[0144] Table 23

[0145]

[0146] From the above results, it can be seen that the separation of each chromatographic peak is better at a column temperature of about 30°C. When the column temperature changes between 28-32°C, the separation of each chromatographic peak is basically the same. This method has good durability for different column temperatures.

[0147] 5. Investigation of different flow rates

[0148] The same sample solution was taken and analyzed at different flow rates of 0.9-1.1 ml / min according to the determination method in Example 1. A chromatogram was obtained, and the relative retention time and relative peak area of ​​each characteristic peak and peak S in the chromatogram were examined. The results of the examination are shown in the following table.

[0149] Table 24

[0150]

[0151]

[0152] Table 25

[0153]

[0154] It can be seen from the above results that when the flow rate changes, the separation degree of peaks 3 and 4 is greatly affected, resulting in a large difference in relative peak areas. Therefore, the present invention preferably sets a fixed flow rate of 1.0 ml per minute as the flow rate for measuring the characteristic spectrum of the granules of the lice formula.

[0155] Example 4

[0156] 1. Investigation of extraction solvent

[0157] Combined with the preparation method of freeze-dried powder, the extraction effects of six extraction solvents including water, ethanol, methanol, 30% methanol, 50% methanol, and 70% methanol were investigated.

[0158] Take about 0.2g of the powder of this product, add 25ml each of water, ethanol, methanol, 30% methanol, 50% methanol, and 70% methanol, respectively, and ultrasonically treat (power 300W, frequency 40kHz) for 30 minutes. Take out, let cool, and then weigh the weight. Make up the lost weight with the corresponding solvent, shake well, filter, and take the filtrate to obtain.

[0159] The survey results are shown in the table below.

[0160] Table 26

[0161]

[0162]

[0163] A comparative study was conducted on different extraction solvents. The extraction effects of water, methanol, ethanol, 30% methanol, 50% methanol, and 70% methanol on the standard decoction of He Li medicinal pieces were poor. The peak shape of the ethanol part was poor, while pure water, 30% methanol, 50% methanol, and 70% methanol could better extract the chromatographic peaks. Among them, 50% methanol had a better peak shape, relatively better system adaptability parameters, and a larger characteristic peak response. Therefore, 50% methanol was selected as the extraction solvent for this experiment.

[0164] 2. Investigation of treatment methods

[0165] The two treatment methods of ultrasonic treatment and heating reflux were compared, and the appropriate extraction method was determined based on the peak information volume and system suitability as the main evaluation indicators.

[0166] Specifically, approximately 0.2 g of the powdered product was added to 25 ml of 50% methanol. The product was then ultrasonically treated (power 300 W, frequency 40 kHz) and heated under reflux for 30 minutes each. The product was removed, allowed to cool, and then weighed. The weight loss was made up with 70% methanol, shaken well, filtered, and the filtrate was obtained. The test results are shown in the following table.

[0167] Table 27

[0168]

[0169] The above results show that the system adaptability parameters of the chromatographic peaks of the test samples of the two extraction methods of the standard decoction of the lice decoction are better, the separation and symmetry are better, and there is no obvious difference in the extraction of the standard decoction of the lice decoction by different extraction methods. Considering the simplicity of operation, ultrasonic treatment is selected as the extraction method of the characteristic spectrum of the standard decoction of the lice decoction.

[0170] 3. Investigation of extraction time

[0171] Take approximately 0.2g of this powder and add 25ml of 50% methanol. Ultrasonicate (power 300W, frequency 40kHz) for 30, 45, and 60 minutes, respectively. Remove, cool, and reweigh. Make up the lost weight with 50% methanol. Shake well, filter, and collect the filtrate. The test results are shown in the following table.

[0172] Table 28

[0173]

[0174] The test results show that when the ultrasound time is 30 to 60 minutes, the peak area of ​​the standard decoction of He Li medicine slices does not differ much, indicating that complete extraction can be achieved within the ultrasound time range of 30 to 60 minutes. Therefore, 30 minutes is used as the ultrasound time for the standard decoction of He Li medicine slices.

[0175] 4. Investigation of different dosages

[0176] Accurately weigh 0.2g of each granule from the lice formula and place it in a stoppered conical flask. Add 15ml, 25ml, and 50ml of 50% methanol, respectively, and weigh the weight. Then, add 0.4g of the granule formula to 25ml of 50% methanol and sonicate (power 300W, frequency 40kHz) for 30 minutes. Let cool, weigh again, and make up the lost weight with 50% methanol. Shake well and filter to obtain the test solution. Accurately pipette 1μL of this solution and inject it into an ultra-high performance liquid chromatograph. The test results are shown in the following table.

[0177] Table 29

[0178]

[0179] When the sampling volume is between 0.1g and 0.4g and the solvent dosage is between 15 and 50ml, the peak area increases proportionally with the increase of dosage and the decrease of solvent dosage, indicating that complete extraction can be achieved when the sampling volume is 0.1g to 0.4g and 15ml to 50ml. Taking the response of the chromatographic peak into consideration, 0.2g / 25ml is used as the sampling volume for the standard decoction of He Li medicinal slices.

[0180] 5. Investigation of injection volume

[0181] Based on the extraction conditions determined above, we further investigated the effect of different injection volumes of the test solution: 5μl, 10μl, 15μl, and 20μl. The test results are shown in the table below.

[0182] Table 30

[0183]

[0184] The test results show that when the injection volume is 5-20μl, no overloading phenomenon occurs. The peak shape of 10μl is better, and the peak height and peak width are relatively moderate. Therefore, the injection volume of 10μl is selected as the injection volume of the test solution in this experiment.

[0185] Obviously, the above embodiments are merely examples for clarity of explanation and are not intended to limit the implementation methods. Those skilled in the art will readily appreciate that other variations or modifications based on the above descriptions are possible. It is not necessary and impossible to enumerate all implementation methods here. Obvious variations or modifications arising therefrom remain within the scope of protection of the present invention.

Claims

1. A method for detecting louse, characterized in that: The method comprises adopting high performance liquid chromatography to detect and obtain a characteristic spectrum, wherein the chromatographic conditions of the high performance liquid chromatography are: Use methanol as mobile phase A and 0.2% phosphoric acid aqueous solution as mobile phase B. Elution was performed according to the following gradient elution program 1, gradient elution program 2, gradient elution program 3, or gradient elution program 5: Gradient elution program 1: Gradient elution program 2: Gradient elution program 3: Gradient elution program 5: The chromatographic column was Tnature C18; The detection wavelength is 240 nm; The preparation of the test solution is as follows: taking the substance to be tested, accurately weighing it, adding a solvent, treating it, cooling it, adding a solvent to make up for the loss, shaking it well, filtering it, and taking the filtrate to obtain the solution; The solvent used in the preparation of the test solution is methanol, water or methanol-water solution; The reference substance was chlorogenic acid.

2. The detection method according to claim 1, wherein Among the chromatographic conditions, the flow rate is 0.9-1.1 ml / min; and / or the column temperature is 28-32° C.; and / or the theoretical plate number calculated based on the chlorogenic acid peak should be no less than 5000.

3. The detection method according to claim 1 or 2, characterized in that The treatment method is heating reflux or ultrasound.

4. The detection method according to claim 3, characterized in that The ultrasonic treatment time is 30-60 min; and / or the ultrasonic power is 250-300 W and the frequency is 40 kHz; And / or, the heating reflux treatment time is 30-60 minutes.

5. The detection method according to claim 1, wherein The added amount of the solvent is 50-250 ml / g.

6. The detection method according to claim 1, characterized in that The injection volume of the test solution is 5-20 μl.

7. The detection method according to claim 1 or 2, characterized in that The chromatographic column has a column length of 250 mm, an inner diameter of 4.6 mm, and a particle size of 5 μm.

8. A quality control method for crane lice based on the detection method according to any one of claims 1 to 7.

9. The quality control method according to claim 8, characterized in that: include: The characteristic spectrum of the test sample obtained by the high performance liquid chromatography method includes at least 8 characteristic peaks; among them, peak 2 is chlorogenic acid; peak 2 is the S peak, and the relative retention time of the remaining characteristic peaks should be within ±10% of the specified value; the specified values ​​of the relative retention times of the 8 characteristic peaks are 0.35, 1.00, 1.04, 1.08, 1.27, 1.59, 1.66, and 2.05, respectively.

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