HPLC (High Performance Liquid Chromatography) fingerprint spectrum detection method of throat clearing tablets

Abstract

The invention belongs to the technical field of quality detection of traditional Chinese medicines, and particularly relates to an HPLC (high performance liquid chromatography) fingerprint spectrum detection method of a Qingyanli tablet, which comprises the step of detecting the Qingyanli tablet by adopting a high performance liquid chromatography, and the chromatographic conditions are as follows: an acetic acid aqueous solution with the mass concentration of 1% is used as a mobile phase A, acetonitrile is used as a mobile phase B, gradient elution is performed, and the HPLC fingerprint spectrum detection method is used for detecting the Qingyanli tablet. According to the method disclosed by the invention, each chromatographic peak is well separated, the characteristic peaks are more, the peak shape is good, the separation degree is better, the detection can be more accurate, the method has better detection stability, consistency and controllability, and the quality of the throat clearing tablets can be more comprehensively and reliably detected.

Description

Technical Field

[0001] The invention belongs to the technical field of quality detection of traditional Chinese medicines, and particularly relates to an HPLC fingerprint detection method for clearing throat and relieving throat tablets. Background Art

[0002] The Qingyanlihou teabag is an empirical prescription of Wei Xiangming, a famous old Chinese medicine doctor from Hunan Institute of Traditional Chinese Medicine. It is composed of raw rehmannia root, ophiopogon japonicus, thunbergii bulb, platycodon grandiflorum, northern adenophora, white peony root, peony bark, mint, scrophularia, licorice, green fruit, and chrysanthemum. It has the effects of nourishing yin, clearing heat, moistening the lungs and relieving cough. The prescription obtained the preparation approval number in 1997 and has been used in clinical practice since then. It has a unique effect in treating chronic pharyngitis. However, since the dosage form is a teabag, it needs to be taken with water and is inconvenient to carry, which is not conducive to long-term medication. As an oral preparation, throat lozenges are easy to carry and take, directly act on the throat mucosa, can quickly relieve symptoms, improve the patient's quality of life, and reduce dependence on other drugs. Therefore, changing the Qingyanlihou teabag to a throat lozenge is of great significance for clinical application.

[0003] The preparation method of the throat-clearing tablets is as follows: the extract is concentrated and dried to obtain a dry paste, the dry paste is combined with crude drug powder and auxiliary materials in proportion, 85% ethanol is used to make a soft material, the particles are dried at 50-60°C, sieved, and magnesium stearate with a mass concentration of 0.5% is added as a lubricant, and tablets are pressed. The extract is extracted by water extraction, and the extracted Chinese medicine formula is: 3.6g of raw rehmannia root, 0.9g of platycodon, 2.7g of white peony root, 2.7g of peony bark, 0.9g of mint, 1.35g of thunbergii, 2.7g of scrophularia, 0.9g of liquorice, 2.7g of northern adenophora, 2.7g of green fruit, 2.7g of ophiopogon, and 1.35g of chrysanthemum. In the prescription, 1 / 2 dose of peony bark and all doses of mint are crushed into fine powder for standby use, which is the crude drug powder. The water extraction process is: 1 / 2 dose of peony bark, with the remaining 10 Chinese medicinal pieces such as raw rehmannia, add 10 times the amount of water for the first time, decoct for 2.0 hours, and add 9 times the amount of water for the second time, decoct for 1.5 hours. The conditions for extract concentration are: temperature 60-70°C, vacuum degree -0.06-0.1Mpa; the conditions for extract drying are: temperature 60-70°C, vacuum degree -0.06-0.1Mpa; the type of auxiliary material is soluble starch, and its dosage ratio is: dry paste powder: raw drug powder: soluble starch = 1: 0.2892: 0.64; the wetting agent is ethanol with a volume concentration of 85%, and the dosage is 30%-35%; a flavoring agent can be added, and the flavoring agent is stevioside with a mass concentration of 1% and menthol with a mass concentration of 0.19%.

[0004] With the increasing application of traditional Chinese medicine, the current single component control can no longer meet the requirements of overall quality control of preparations. The characteristic spectrum can reflect the chemical composition and distribution characteristics of traditional Chinese medicine, as well as its distribution ratio, content characteristics, potential physiological activity and quantitative characteristics of efficacy, which is consistent with the integrity, characteristics, multi-target and nonlinearity that should be reflected in the quality control of traditional Chinese medicine. Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide an HPLC fingerprint detection method for Qingyan Lihou tablets to improve the accuracy of quality detection of Qingyan Lihou tablets.

[0006] The embodiment of the present invention provides an HPLC fingerprint detection method for Qingyan Lihou Tablets, comprising detecting Qingyan Lihou Tablets by high performance liquid chromatography, wherein the chromatographic conditions include: using 1% mass concentration of acetic acid aqueous solution as mobile phase A, using acetonitrile as mobile phase B, gradient elution, and the procedure of gradient elution is:

[0007]

[0008] Preferably, in the chromatographic conditions, the flow rate is 1.0 mL / min.

[0009] Preferably, the column temperature is 30°C.

[0010] Preferably, the detection wavelength is 254 nm.

[0011] Preferably, in the chromatographic conditions, the chromatographic column is Ultimate XB-C18.

[0012] Preferably, during the detection, the preparation method of the test solution is to take the Qingyan Lihou tablets, grind them, add methanol, perform ultrasonic treatment, and filter to obtain the test solution.

[0013] Preferably, the weight volume of the throat clearing and throat relieving tablets and methanol is 2g:25mL.

[0014] Preferably, the power of ultrasonic treatment is 300 W and the frequency is 40 kHz.

[0015] Preferably, the method further comprises high performance liquid chromatography detection of a reference substance solution, wherein the reference substances are gallic acid, chlorogenic acid, paeoniflorin, liquiritin and paeonol.

[0016] The beneficial effect of the present invention is that the present invention has found through experiments that when high performance liquid chromatography is used to detect the Qingyan Lihou tablets, an acetic acid aqueous solution with a mass concentration of 1% is selected as the mobile phase A, and acetonitrile is selected as the mobile phase B, and the gradient elution procedure as described in the present application is performed for gradient elution. Each chromatographic peak can be well separated, and there are more characteristic peaks, good peak shape, and good separation degree. The detection can be more accurate, and the detection has good detection stability, consistency and controllability, and the quality of the Qingyan Lihou tablets can be detected more comprehensively and reliably. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 This is the characteristic spectrum of Example 1 of the present invention.

[0018] Figure 2 It is the characteristic spectrum of comparative example 1 of the present invention, Figure 3 It is the characteristic spectrum of comparative example 2 of the present invention, Figure 4 It is the characteristic spectrum of comparative example 3 of the present invention, Figure 5 It is the characteristic spectrum of comparative example 4 of the present invention, Figure 6 It is the characteristic spectrum of comparative example 5 of the present invention, Figure 7 It is the characteristic spectrum of comparative example 6 of the present invention, Figure 8 It is the characteristic spectrum of comparative example 7 of the present invention, Fig. 9 It is the characteristic spectrum of comparative example 8 of the present invention, Fig.10 It is the characteristic spectrum of comparative example 9 of the present invention, Fig.11 is the characteristic spectrum of comparative example 10 of the present invention, Fig.12 is the characteristic spectrum of comparative example 11 of the present invention, Fig.13 It is the characteristic spectrum of comparative example 12 of the present invention.

[0019] Fig.14 is the characteristic spectrum of Example 2 at a wavelength of 254 nm, Fig.15 is the characteristic spectrum of Example 2 at a wavelength of 327 nm, Fig.16 is the characteristic spectrum of Example 2 at a wavelength of 210 nm, Fig.17 is the characteristic spectrum of Example 2 at a wavelength of 230 nm, Fig.18 This is the characteristic spectrum of Example 2 at a wavelength of 280 nm.

[0020] Fig.19 This is the characteristic spectrum of the extraction solvent investigated in Example 3.

[0021] Fig. 20 The characteristic spectrum of the test solution of Example 4 is as follows: Fig.21 is the characteristic spectrum of the detection reference solution of Example 4, Fig. 22 This is the characteristic spectrum of the blank group solution of Example 4.

[0022] Fig.23 This is the chromatogram of the double negative sample lacking white peony root and peony bark in Example 4, Fig.24 The negative sample chromatogram of the lack of white peony root of Example 4 is shown in FIG. Fig.25 This is the chromatogram of the negative sample lacking Paeonia suffruticosa root bark of Example 4, Fig.26 The chromatogram of the negative sample lacking chrysanthemum in Example 4 is shown in FIG. Fig. 27 This is the chromatogram of the negative sample lacking licorice in Example 4, Fig.28 The chromatogram of the negative sample lacking Scrophularia ningpoensis of Example 4 is shown in FIG. Fig.29 is the negative sample chromatogram of Rehmannia glutinosa of Example 4, Fig.30 This is the chromatogram of the green fruit negative sample of Example 4, Fig.31 This is the chromatogram of the negative sample lacking Glehnia littoralis of Example 4, Fig.32 This is the chromatogram of the mint-deficient negative sample of Example 4, Fig.33 This is the attribution of the characteristic spectrum peaks of Example 4. DETAILED DESCRIPTION

[0023] Instruments and reagents

[0024] (1) Instrument

[0025] Agilent Technologies 1260 InfinityⅡ high performance liquid chromatograph, DAD detector; SB-5200D ultrasonic cleaner (Ningbo Xinzhi Biotechnology Co., Ltd.); Lingsheng multifunctional crusher (Yongkang Hongtaiyang Electromechanical Co., Ltd.); ES-502HA electronic balance (Changsha Xiangping Technology Development Co., Ltd.); SQP electronic analytical balance (Sartorius Scientific Instruments (Beijing) Co., Ltd.); XPE105 electronic analytical balance (METTLER).

[0026] (2) Drug testing

[0027] The preparation method of the throat-clearing and throat-relief tablets is as follows: the extract is concentrated and dried to obtain a dry paste, the dry paste is combined with crude drug powder and auxiliary materials in proportion, 85% ethanol is used to make a soft material, the particles are dried at 50-60°C, sieved, 0.5% magnesium stearate is added as a lubricant, and tablets are pressed.

[0028] The extract is extracted by water, and the extracted Chinese medicine formula is: 3.6g of raw rehmannia root, 0.9g of platycodon grandiflorum, 2.7g of white peony root, 2.7g of peony bark, 0.9g of mint, 1.35g of thunbergii, 2.7g of scrophulariae, 0.9g of licorice, 2.7g of northern adenophora, 2.7g of green fruit, 2.7g of ophiopogon japonicus, and 1.35g of chrysanthemum. In the prescription, 1 / 2 dose of peony bark and all doses of mint are crushed into fine powder for standby use, which is the raw medicine powder. The water extraction process is: 1 / 2 dose of peony bark, with the remaining 10 Chinese medicine pieces such as raw rehmannia root, add 10 times the amount of water for the first time, decoct for 2.0h, and add 9 times the amount of water for the second time, decoct for 1.5h. The conditions for extract concentration are: temperature 60-70°C, vacuum degree -0.06-0.1Mpa; the conditions for extract drying are: temperature 60-70°C, vacuum degree -0.06-0.1Mpa; the type of auxiliary material is soluble starch, and its usage ratio is: dry paste powder: crude drug powder: soluble starch = 1:0.2892:0.64; ethanol with a volume concentration of 85% is used as a wetting agent, and the usage is 30%-35%.

[0029] Acetonitrile and methanol were of chromatographic grade, water was purified water, and other reagents were of analytical grade.

[0030] Reagents: chromatographic methanol (Sigma-Aldrich, 99.9%), chromatographic acetonitrile (Sigma-Aldrich, 99.9%), purified water (Yibao purified water), analytical methanol (Sinopharm Chemical Reagent Co., Ltd.), analytical ethanol (Sinopharm Chemical Reagent Co., Ltd.), phosphoric acid (Shanghai MacLean Biochemical Technology Co., Ltd., 85-90%), glacial acetic acid (Shanghai MacLean Biochemical Technology Co., Ltd., 99.9%).

[0031] Reference substances: paeoniflorin (110736-202145, 99.9%, China Food and Drug Inspection Institutes), paeonol (110708-201407, 99.9%, China Food and Drug Inspection Institutes), chlorogenic acid (110753-201817, 96.8%, China Food and Drug Inspection Institutes), glycyrrhizin (111610-202209, 95.2%, China Food and Drug Inspection Institutes), gallic acid (110831-201906, 91.5%, China Food and Drug Inspection Institutes).

[0032] Example 1

[0033] A HPLC fingerprint detection method for Qingyan Lihou Tablets is used to detect Qingyan Lihou Tablets by high performance liquid chromatography, wherein the chromatographic conditions include: Ultimate XB-C18 (4.6×250mm, 5μm) chromatographic column; flow rate 1.0mL / min, column temperature 30°C, 1% mass concentration glacial acetic acid solution as mobile phase A, acetonitrile as mobile phase B; gradient elution according to the provisions in the following table, washing and pre-equilibration for 35min. The detection wavelength is 254nm, and the gradient elution program is shown in Table 1.

[0034] Table 1 Gradient elution program of characteristic spectrum of Qingyan Lihou tablets

[0035]

[0036] The preparation method of the test solution is as follows: take an appropriate amount of Qingyan Lihou Tablets, grind them into powder, take about 2g, accurately weigh, place in a stoppered conical flask, accurately add 25ml of methanol, ultrasonically treat (power 300W, frequency 40kHz) for 30 minutes, take out and cool, take the supernatant and pass it through a microporous filter to obtain the test solution.

[0037] Determination method: Accurately draw 10 μL of the test solution, inject it into the liquid chromatograph, measure, and record the chromatogram to obtain the result. Figure 1 As shown,

[0038] The experimental results are as follows Figure 1 As shown, the results show that the separation of each chromatographic peak is good.

[0039] Comparative Example 1

[0040] The preparation method of the test solution is as follows: take an appropriate amount of Qingyan Lihou Tablets, grind them into powder and sieve them (80 mesh), take about 2.0 g, accurately weigh, place in a stoppered conical flask, accurately add 25 ml of pure methanol, seal it, weigh it, ultrasonically treat it (power 300 W, frequency 40 kHz) for 30 minutes, cool it, weigh it again, make up the lost weight with pure methanol, shake it well, filter it, and take the filtrate to obtain it.

[0041] The Qingyan Lihou Tablets were detected by high performance liquid chromatography, and the chromatographic conditions were: Ultimate XB-C18 (4.6×250mm, 5μm) chromatographic column, flow rate 1.0mL / min, column temperature 30°C, wavelength 256nm, injection volume 10μl, 0.5% mass concentration phosphoric acid solution as mobile phase A, acetonitrile as mobile phase B, and gradient elution according to the provisions in Table 2 below.

[0042] Table 2 Characteristic spectrum of Qingyan Lihou tablets Gradient elution program 1

[0043]

[0044] The experimental results are as follows Figure 2 As shown, the results show that the chromatographic peaks are dense in the time period of 36-44 minutes and the separation is insufficient.

[0045] Comparative Example 2

[0046] The difference between Comparative Example 2 and Comparative Example 1 is that the elution procedure is different, and the other procedures are the same as Comparative Example 1. The elution procedure of Comparative Example 2 is shown in Table 3.

[0047] Table 3 throat clearing characteristic spectrum gradient elution program 2

[0048]

[0049] The experimental results are as follows Figure 3 As shown, the results show that the chromatographic peaks are dense in the 22-30min period and the separation is insufficient.

[0050] Comparative Example 3

[0051] Comparative Example 3 is different from Comparative Example 1 in that the elution procedure is different, and the other procedures are the same as Comparative Example 1. The elution procedure of Comparative Example 3 is shown in Table 4.

[0052] Table 4 Throat-clearing characteristic spectrum gradient elution program three

[0053]

[0054] The experimental results are as follows Figure 4 As shown, the results show that the chromatographic peaks are dense in the 44-50min period and the separation is insufficient.

[0055] Comparative Example 4

[0056] The preparation method of the test solution of Comparative Example 4 is the same as that of Example 1.

[0057] The Qingyan Lihou Tablets were detected by high performance liquid chromatography, and the chromatographic conditions were: Ultimate XB-C18 (4.6×250mm, 5μm) chromatographic column, flow rate 1.0mL / min, column temperature 30°C, wavelength 254nm, injection volume 10μl, 1% mass concentration glacial acetic acid solution as mobile phase A, acetonitrile as mobile phase B, and gradient elution according to the provisions in Table 5.

[0058] Table 5 throat clearing characteristic spectrum gradient elution program four

[0059]

[0060] The experimental results are as follows Figure 5 As shown, the results show that the chromatographic peaks are scarce.

[0061] Comparative Example 5

[0062] Comparative Example 5 is different from Comparative Example 4 in that the elution procedure is different, and the other procedures are the same as Comparative Example 4. The elution procedure of Comparative Example 5 is shown in Table 6.

[0063] Table 6 Throat Clearing Characteristic Spectrum Gradient Elution Program 5

[0064]

[0065] The experimental results are as follows Figure 6 As shown, the results show that some chromatographic peaks overlap.

[0066] Comparative Example 6

[0067] Comparative Example 6 is different from Comparative Example 4 in that the elution procedure is different, and the other procedures are the same as Comparative Example 4. The elution procedure of Comparative Example 6 is shown in Table 7.

[0068] Table 7 Throat Clearing Characteristic Spectrum Gradient Elution Program 7

[0069]

[0070] The experimental results are as follows Figure 7 As shown, the results show that the chromatographic peaks are denser in the 65-75min period, which is not as good as program six.

[0071] Comparative Example 7

[0072] Comparative Example 7 is different from Comparative Example 4 in that the elution procedure is different, and the other procedures are the same as Comparative Example 4. The elution procedure of Comparative Example 7 is shown in Table 8.

[0073] Table 8 throat clearing characteristic spectrum gradient elution program eight

[0074]

[0075]

[0076] The experimental results are as follows Figure 8 As shown, the results show that the chromatographic peaks are basically good.

[0077] Comparative Example 8

[0078] The preparation method of the test solution of Comparative Example 8 is the same as that of Comparative Example 7.

[0079] The Qingyan Lihou Tablets were detected by high performance liquid chromatography, and the chromatographic conditions were: ShimNex CS C18 column (250 mm×4.6 mm, 5 μm), flow rate 1.0 mL / min, column temperature 30°C, wavelength 254 nm, injection volume 10 μl, 1% glacial acetic acid solution as mobile phase A, acetonitrile as mobile phase B, and gradient elution as specified in the table below.

[0080] Table 9 throat clearing characteristic spectrum gradient elution program six

[0081]

[0082] The experimental results are as follows Fig. 9 As shown, the results show that the chromatographic peaks are basically good, but the peak shape is not as good as the Ultimate XB-C18 column.

[0083] Comparative Example 9

[0084] Comparative Example 9 is different from Comparative Example 8 in that the elution procedure is different, and the other procedures are the same as Comparative Example 8. The elution procedure of Comparative Example 9 is shown in Table 10.

[0085] Table 10 Throat Clearing Characteristic Spectrum Gradient Elution Program Nine

[0086]

[0087] The experimental results are as follows Fig.10 As shown, the results show that the chromatographic peaks are not good.

[0088] Comparative Example 10

[0089] Comparative Example 10 is different from Comparative Example 8 in that the elution procedure is different, and the other procedures are the same as Comparative Example 8. The elution procedure of Comparative Example 10 is shown in Table 11.

[0090] Table 11 Throat Clearing Characteristic Spectrum Gradient Elution Program 10

[0091]

[0092] The experimental results are as follows Fig.11 As shown, the results show that the chromatographic peaks are not good.

[0093] Comparative Example 11

[0094] Comparative Example 11 is different from Comparative Example 8 in that the elution procedure is different, and the other procedures are the same as Comparative Example 8. The elution procedure of Comparative Example 11 is shown in Table 12.

[0095] Table 12 Throat Clearing Characteristic Spectrum Gradient Elution Program 11

[0096]

[0097] The experimental results are as follows Fig.12 As shown, the results show that the chromatographic peak is not good, and further adjustment of the elution gradient is attempted.

[0098] Comparative Example 12

[0099] Comparative Example 12 is different from Comparative Example 8 in that the elution procedure is different, and the other procedures are the same as Comparative Example 8. The elution procedure of Comparative Example 12 is shown in Table 13.

[0100] Table 13 throat clearing characteristic spectrum gradient elution program twelve

[0101]

[0102] The experimental results are as follows Fig.13 As shown, the results show that the chromatographic peaks are not good.

[0103] Example 2

[0104] Wavelength selection

[0105] Compared with Example 1, the difference between Example 2 and Example 1 is the wavelength, and the other aspects are the same as Example 1. The wavelengths are respectively: 210nm, 230nm, 254nm, 280nm, and 327nm.

[0106] Compare the chromatograms at 210nm, 230nm, 254nm, 280nm, and 327nm. The results are shown in Figure 14-18 .

[0107] The results show that the baseline is not stable at 210nm and 230nm wavelengths, and the absorption signals of peaks above 280nm are reduced. However, the peaks are relatively well separated at 254nm and 327nm wavelengths, and more characteristic peaks are obtained with better overall peak shape. However, the later standard is best at 254nm, so 254nm is selected as the wavelength.

[0108] Example 3

[0109] Investigation of the preparation method of test solution

[0110] Extraction solvent

[0111] Take an appropriate amount of this product, take 2g, a total of 3 portions, accurately weigh, put in a stoppered conical flask, accurately add 25ml of methanol, ethanol, and distilled water respectively, treat with ultrasound (power 300W, frequency 40kHz) for 30 minutes, take out and cool, take the supernatant and pass it through a 0.22μm microporous filter head to obtain.

[0112] Other conditions are the same as in Example 1. The experimental results are shown in Fig.19 .

[0113] During sample treatment, water ultrasonic treatment of samples resulted in slower filtration. The number and area of ​​peaks in samples treated with water and ethanol were significantly smaller than those in samples extracted with methanol.

[0114] Example 4

[0115] Methodological validation

[0116] (1) Chromatographic peak identification

[0117] According to the HPLC fingerprint detection method of Example 1, the test solution, reference solution (gallic acid, chlorogenic acid, paeoniflorin, liquiritin, paeonol) and solvent blank (methanol) of Example 1 were detected in turn. The preparation method of the reference solution is as follows: accurately weigh appropriate amounts of paeoniflorin, paeonol, chlorogenic acid, liquiritin and gallic acid reference, add methanol to prepare a solution containing 25.10 μg·mL of paeoniflorin. -1 , Paeonol 28.51 μg·mL -1 , chlorogenic acid 26.54 μg·mL -1 , Liquorice glycoside 20.30μg mL -1 and gallic acid 22.73 μg·mL -1 of mixed reference solution.

[0118] Results Figure 20-22 .

[0119] By comparing the peak time, spectra of the reference and test samples, it was determined that peak 1 marked in the figure was gallic acid, peak 3 was chlorogenic acid, peak 7 was paeoniflorin, peak 9 was liquiritigenin and peak 16 was paeonol, and the peak shape and separation were relatively good.

[0120] (2) Peak attribution

[0121] According to the test sample preparation method of Example 1, 10 negative samples (the double negative sample lacking white peony root and moutan bark, the negative sample lacking white peony root, the negative sample lacking moutan bark, the negative sample lacking chrysanthemum, the negative sample lacking liquorice, the negative sample lacking Scrophulariaceae, the negative sample lacking Rehmannia glutinosa, the negative sample lacking green fruit, the negative sample lacking Adenophora adenophora, and the negative sample lacking Mentha sutchuenensis) and the test sample were processed.

[0122] According to the HPLC fingerprint detection method of Example 1, the samples were injected and tested respectively, and the results are shown in Figure 23-33 .

[0123] The results showed that 8 chromatographic peaks, including 2, 5, 9, 10, 13, 15 and 16, were temporarily selected as common peaks. Among them, peaks 2, 5 and 9 belonged to Qingguo, peaks 10 and 13 belonged to Chrysanthemum, peak 15 belonged to Mentha, and peak 16 belonged to Paeonia suffruticosa. There were no peaks for Scrophularia and Rehmannia glutinosa.

[0124] (3) Precision test

[0125] According to the determined analytical method, one sample solution was prepared and injected 6 times continuously to examine the precision. Eight characteristic peaks were calibrated, and peak 16 (paeonol) was used as the reference peak. The relative retention time and relative peak area of ​​other characteristic peaks were calculated. The results are shown in Tables 14-15.

[0126] Table 14 Characteristic spectrum precision relative retention time results

[0127]

[0128]

[0129] Table 15 Characteristic spectrum precision relative peak area results

[0130]

[0131] The results showed that the relative retention time RSD of the eight common peaks did not exceed 1.0%, and the relative peak area RSD did not exceed 5.0%, indicating that the instrument had good precision.

[0132] (4) Repeatability test

[0133] According to the determined analytical method, 6 test solutions were prepared and tested in sequence to examine the repeatability. 12 characteristic peaks were calibrated, with peak 16 (paeonol) as the reference peak, and the relative retention time and relative peak area of ​​other characteristic peaks were calculated. The results are shown in Table 16-17.

[0134] Table 16 Characteristic spectrum repeatability relative retention time results

[0135]

[0136] Table 17 Characteristic spectrum repeatability relative peak area results

[0137]

[0138] The results showed that the relative retention time RSD of the eight common peaks did not exceed 1.0%, and the relative peak area RSD did not exceed 5.0%, indicating that the method had good repeatability.

[0139] (5) Solution stability test

[0140] According to the determined analytical method, one sample solution was prepared, and the characteristic spectrum was measured at 0h, 2h, 4h, 6h, 8h, 12h, and 24h to examine the stability. Eight characteristic peaks were calibrated, and peak 16 (paeonol) was used as the reference peak. The relative retention time and relative peak area of ​​other characteristic peaks were calculated. The results are shown in Tables 18 to 19.

[0141] Table 18 Characteristic spectrum stability relative retention time results

[0142]

[0143] Table 19 Characteristic spectrum stability relative peak area results

[0144]

[0145] The results showed that the relative retention time RSD of the eight common peaks did not exceed 1.0%, and the relative peak area RSD did not exceed 5.0%, indicating that the solution had good stability within 24 hours.

[0146] A person skilled in the art should understand that the discussion of any of the above embodiments is merely illustrative and is not intended to imply that the scope of protection of the present application is limited to these examples. In line with the concept of the present application, the technical features in the above embodiments or different embodiments may be combined, the steps may be implemented in any order, and there are many other variations of different aspects of one or more embodiments of the present application as described above, which are not provided in detail for the sake of simplicity.

[0147] One or more embodiments of the present application are intended to cover all such substitutions, modifications and variations that fall within the broad scope of the present application. Therefore, any omissions, modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of one or more embodiments of the present application should be included in the protection scope of the present application.

Claims

1. A HPLC fingerprint detection method for throat clearing tablets, characterized in that: The method comprises using high performance liquid chromatography to detect the throat clearing tablets, wherein the chromatographic conditions include: using 1% acetic acid aqueous solution as mobile phase A, using acetonitrile as mobile phase B, and gradient elution, and the procedure of the gradient elution is:

2. The HPLC fingerprint detection method of the Qingyan Lihou Tablets as claimed in claim 1, characterized in that: In the chromatographic conditions, the flow rate was 1.0 mL / min.

3. The HPLC fingerprint detection method of the Qingyan Lihou Tablets as claimed in claim 1 is characterized in that: The column temperature was 30°C.

4. The HPLC fingerprint detection method of the throat clearing tablets as claimed in claim 1 is characterized in that: The detection wavelength is 254 nm.

5. The HPLC fingerprint detection method of the throat clearing tablets as claimed in claim 1 is characterized in that: In the chromatographic conditions, the chromatographic column was Ultimate XB-C18.

6. The HPLC fingerprint detection method of the throat clearing tablets as claimed in claim 1 is characterized in that: During the test, the method for preparing the test solution is to take the Qingyan Lihou tablets, grind them, add methanol, perform ultrasonic treatment, and filter to obtain the test solution.

7. The HPLC fingerprint detection method of the throat clearing tablets as claimed in claim 6 is characterized in that: The weight volume of the throat clearing and throat relieving tablets and methanol is 2g:25mL.

8. The HPLC fingerprint detection method of the throat clearing tablets as claimed in claim 6 is characterized in that: The power of ultrasonic treatment was 300 W and the frequency was 40 kHz.

9. The HPLC fingerprint detection method of the Qingyan Lihou Tablets according to any one of claims 1 to 8, characterized in that: The method also includes high performance liquid chromatography detection of reference substance solutions, wherein the reference substances are gallic acid, chlorogenic acid, paeoniflorin, liquiritin and paeonol.

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