CN115343395B - Establishment method of UPLC fingerprint of scattered decoction and standard fingerprint and application thereof - Google Patents

Establishment method of UPLC fingerprint of scattered decoction and standard fingerprint and application thereof Download PDF

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CN115343395B
CN115343395B CN202211035298.0A CN202211035298A CN115343395B CN 115343395 B CN115343395 B CN 115343395B CN 202211035298 A CN202211035298 A CN 202211035298A CN 115343395 B CN115343395 B CN 115343395B
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王龙虎
李丽平
顾选
范玛莉
赵帅丹
王莎
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Huayi Pharmaceutical Co Ltd
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Abstract

The application relates to the field of traditional Chinese medicines, and in particular discloses a method for establishing a UPLC fingerprint of a scattered decoction, a standard fingerprint and application thereof, wherein the method for establishing the UPLC fingerprint comprises the following steps: (a) preparing a control solution; (b) Adding methanol solution into the decoction, weighing, ultrasonic treating, cooling, weighing again, supplementing the weight with methanol solution, mixing, and filtering to obtain filtrate as sample solution; (c) Sucking a reference solution and a sample solution, and collecting a chromatogram; (d) Selecting chromatographic peaks existing in different batches of scattered chromatograms as common peaks to obtain standard fingerprint of scattered decoction; the application also discloses the standard fingerprint spectrum of the yao-decoction UPLC obtained by the method and the application of the standard fingerprint spectrum in the detection of the yao-decoction preparation. The obtained UPLC fingerprint can rapidly and comprehensively reflect and control the overall quality problem of the scattered soup, and the quality control of enterprises on the scattered soup products is met.

Description

Establishment method of UPLC fingerprint of scattered decoction and standard fingerprint and application thereof
Technical Field
The application relates to the technical field of traditional Chinese medicines, in particular to a method for establishing a UPLC fingerprint of a yaoyan decoction, and a standard fingerprint and application thereof.
Background
Migraine is a common neurovascular disease, is mainly a moderately severe pulsating headache repeatedly attacks on one side or two sides of the temporal part, can be accompanied by vision and body premonitory symptoms before attack, is frequently accompanied by vomiting, is co-morbid with various diseases such as depression, anxiety, insomnia, cognitive dysfunction and the like, can cause various complications such as leukoencephalopathy, cognitive decline, asymptomatic cerebral infarction and the like, and seriously jeopardizes the physical health and life quality of people.
The western medicine has single action mechanism, has insignificant treatment effect on migraine, and can not comprehensively regulate the clinical treatment of migraine, so that the unique clinical dialectical treatment system of the traditional Chinese medicine receives more and more attention. The ancient famous prescription bias-dispelling decoction contains eight traditional Chinese medicinal materials including ligusticum wallichii, liquorice, radix angelicae, radix paeoniae alba, semen pruni, radix bupleuri, rhizoma cyperi and semen brassicae. According to hundreds of years of actual Linchuan application and scientific research, the migraine-relieving decoction has obvious curative effect on migraine.
In 2018, 4 months is the implementation of the Chinese medicine law of the people's republic of China, the stable development of the Chinese medicine compound preparation from the ancient classical prescription is promoted, better guarantee is provided for the health of people, and the national Chinese medicine administration and the national drug administration make the catalog (first batch) of the ancient classical prescription, wherein the scattered soup is catalog 98. However, the report of the off-set decoction is mainly focused on the aspects of clinical medication research and pharmacological research, and the related research on the quality control of the product detection is less.
And because the ingredients of the off-set soup are complex and various, the difficulty in controlling the quality of the off-set soup is also improved. The fingerprint is an effective means for comprehensively analyzing the authenticity of traditional Chinese medicine or Chinese patent medicine by using modern analysis technology to evaluate the consistency and stability of the quality of raw material medicine, semi-finished product and finished product, and compared with the existing identification of single effective component or index component, the traditional Chinese medicine fingerprint technology integrates various effective chemical information, so that the result is more scientific and reasonable. At present, the fingerprint of the traditional Chinese medicine becomes an important method for controlling the quality of the traditional Chinese medicine accepted at home and abroad, and the research and establishment of the fingerprint have important significance for improving the quality of the traditional Chinese medicine and promoting the modernization of the traditional Chinese medicine.
The research of the fingerprint establishment of the off-set decoction has important significance for comprehensively and comprehensively reflecting the quality difference of the off-set decoction products and effectively controlling the quality of the whole process. Mo Yujia, wang Yan, etc. establishment of HPLC fingerprint of classical famous prescription off-set decoction and magnitude transfer study of Ligusticum wallichii (J.) Chinese traditional Chinese medicine impurity, 2020.45 (3): 573 and Liu Xuechun, mo Yujia, etc. substance benchmark quantity value transfer analysis (J) of classical formula off-set soup, chinese traditional medicine impurity, 2022.47 (8): 2099 is a reference to the fingerprint of the decoction, but is an HPLC fingerprint method, and the decoction has a large number of components and is complex, and two fingerprints are required to be established by the fingerprint method, which takes much time. Compared with HPLC, the UPLC fingerprint has higher speed, sensitivity and separation degree, so that the establishment of the fingerprint for the yawing decoction by using UPLC has important significance for quality control analysis of the yawing decoction.
Disclosure of Invention
In order to establish a fingerprint of the yao-decoction by adopting UPLC, reflect and control the overall quality of the yao-decoction, the application provides a method for establishing the UPLC fingerprint of the yao-decoction, and a standard fingerprint and application thereof.
In a first aspect, the present application provides a method for establishing a fingerprint of a yao decoction UPLC, which adopts the following technical scheme:
a method for establishing a UPLC fingerprint of a scattered soup comprises the following steps:
(a) Preparation of control solution: taking paeoniflorin reference substance, adding methanol solution to prepare reference substance solution;
(b) Preparation of test solution: adding methanol solution into the decoction, weighing, ultrasonic treating for 30min, taking out, cooling, weighing again, supplementing the reduced weight with methanol solution, mixing, and filtering to obtain filtrate as sample solution;
(c) And (3) measuring: precisely sucking the reference solution and the sample solution respectively, injecting into an ultra-high performance liquid chromatograph, and collecting chromatograms;
(d) Introducing the integral signal of the fingerprint of the test sample solution of the scattered decoction obtained in the step (c) into software of a traditional Chinese medicine chromatographic fingerprint similarity evaluation system, and selecting chromatographic peaks existing in different batches of scattered chromatograms as common peaks to obtain a standard fingerprint of the scattered decoction;
wherein in step (c), the liquid chromatography conditions are: octadecylsilane chemically bonded silica is used as a filler, acetonitrile is used as a mobile phase A, and phosphoric acid solution is used as a mobile phase B; gradient elution: 0-8min, A2→8%;8-15min, A8%;15-25min, A8-15%; 25-27min, A15-20%; 27-37min, A20→25%;37-60min, A25→70%; the detection wavelength was 230nm.
By adopting the technical scheme, the method prepares the fingerprint through preprocessing the mobile phase composition, the gradient elution program, the flow speed, the detection wavelength and the solution of the test sample, performs UPLC fingerprint similarity analysis on 30 batches of scattered soup by using the traditional Chinese medicine chromatographic fingerprint similarity evaluation system, and has the similarity of more than 0.90, so that the method for obtaining the scattered soup UPLC fingerprint is reliable, is simple and convenient to operate, can scientifically and effectively evaluate the quality of the scattered soup, and meets the requirements of enterprises on quality control of scattered soup products;
the relative retention time of the common peaks of all batches of the scattered soup in the fingerprint established by the method is basically consistent, the analysis method is good in stability, the effective components of the scattered soup can be measured, and the finally obtained scattered soup UPLC fingerprint detection method can be used for quality control of the scattered soup to be evaluated and is an effective means for comprehensively evaluating the quality of the scattered soup. According to the method for detecting the UPLC fingerprint of the obtained scattered soup, paeoniflorin is used as a reference peak, UPLC fingerprint of 30 batches of scattered soup is established, 13 common peaks are obtained, analysis and identification are carried out on the obtained UPLC common peaks, and components such as sinapine, ferulic acid, paeoniflorin, albiflorin and glycyrrhizic acid are identified, so that the chemical substance basis of the scattered soup is further elucidated.
Optionally, in step (b), the ultrasonic treatment parameters are: the power was 320w and the frequency was 40kHz.
By adopting the technical scheme, the fingerprint study of the scattered soup by using the UPLC technology is realized through ultrasonic treatment after methanol dissolution of the scattered soup product, and the scattered soup is used as an integral study method of the scattered soup, so that more peaks are generated under the condition, the reaction information is more comprehensive, and the precision is high, and the stability and the repeatability are good.
Optionally, in the step (d), the similarity between the fingerprint of the sample and the fingerprint of the reference sample is not lower than 0.90.
Optionally, 13 peaks are shared, wherein the number of peaks is 13, the number of peaks is sinapine, the number of peaks is paeoniflorin, the number of peaks is reference peak paeoniflorin, the number of peaks is ferulic acid, the number of peaks is glycyrrhizin, the number of peaks is isoliquiritigenin, the number of peaks is benzoylpaeoniflorin, and the number of peaks is glycyrrhizic acid.
Optionally, in the standard fingerprint, paeoniflorin is used as a reference peak, and the relative retention time of the obtained common peak is as follows: peak 1:0.280; peak 2:0.797; peak 3:0.897; peak 4:1.000; peak 5:1.055; peak 6:1.110; peak 7:1.211; peak 8:1.231; peak 9:1.262; peak 10:1.510; peak 11:1.822; peak 12:2.038; peak 13:2.126.
optionally, in the standard fingerprint, paeoniflorin is used as a reference peak, and the relative peak area of the obtained common peak is as follows: peak 1:0.222; peak 2:0.442; peak 3:0.162; peak 4:1.000; peak 5:0.347; peak 6:0.208; peak 7:0.109; peak 8:0.064; peak 9:0.042; peak 10:0.041; peak 11:0.060; peak 12:0.036; peak 13:0.046.
optionally, the bulk soup product in step (b) is prepared from the following raw materials: 777g of white peony root, 1554g of szechuan lovage rhizome, 155g of bunge cherry seed, 155g of bupleurum root, 466g of white mustard seed, 311g of nutgrass galingale rhizome, 155g of liquorice and 78g of dahurian angelica root.
By adopting the technical scheme, the decoction for dispersing the partial deviation has the compatibility characteristics of the ligusticum wallichii for dispelling wind, relieving pain, promoting qi and activating blood, radix bupleuri for benefiting liver and gall, radix paeoniae alba for softening liver and relieving pain and bunge cherry seed with settled drug property. Rhizoma Ligustici Chuanxiong is used for dispelling pathogenic wind and relieving pain, and dispelling pathogenic wind evil outside, entering liver and gall channel to promote qi circulation and blood circulation, and internal qi movement, and is used as monarch drug; radix bupleuri and rhizoma cyperi can dredge liver and gall, and relieve shaoyang, radix angelicae can dispel cold and relieve pain, semen brassicae can dredge qi and resolve phlegm, dredge collaterals and relieve pain, radix paeoniae alba and radix glycyrrhizae can relieve spasm and pain, and the radix angelicae can be used as ministerial drugs; the bunge cherry seed has the effects of promoting urination and defecation, has the main property of descending, can prevent excessive dispersion of the szechuan lovage rhizome Xin Wensheng, and has the wonderful ascending and descending phase with the szechuan lovage rhizome, and is an adjuvant drug; licorice root, radix Glycyrrhizae regulates the action of other drugs as a guiding drug. The medicines are applied together to dispel wind and relieve pain, and liver and gallbladder are good formulas for treating migraine caused by pathogenic wind attacking shaoyang.
Optionally, the preparation of the polarization dispersion soup in the step (b) is prepared by the following method:
weighing 777g of white peony root, 1554g of szechuan lovage rhizome, 155g of bunge cherry seed, 155g of Chinese thorowax root, 466g of white mustard seed, 311g of nutgrass galingale rhizome, 155g of liquoric root and 78g of dahurian angelica root, adding 8 times of water of the total mass, soaking for 30min, boiling, decocting for 30min, filtering, adding 6 times of water into filter residues, boiling, decocting for 25min, filtering, combining the two decoctions, concentrating under reduced pressure at 65 ℃ until the relative density is 1.05-1.10, adding dextrin, spray-drying into dry paste powder, adding dextrin, uniformly mixing, granulating for 1000g, and obtaining the biased soup granule preparation.
By adopting the technical scheme, the decoction for eliminating the offset is prepared into particles, so that the decoction is more convenient to carry and take.
In a second aspect, the application provides a standard fingerprint spectrum of a scattered soup UPLC, which adopts the following technical scheme:
a standard fingerprint of a decoction for treating hypertension, and its preparation method are provided.
In a third aspect, the application provides an application of a standard fingerprint spectrum of a bias-dispersion soup UPLC, which adopts the following technical scheme:
an application of a standard fingerprint spectrum of a yao-decoction UPLC in the detection of a yao-decoction preparation is provided.
By adopting the technical scheme, the standard fingerprint obtained by the method can be used as a control method for controlling the quality of the off-set soup, and can be applied to the identification of the authenticity of the off-set soup and the detection of components. When the method is applied, the fingerprint spectrum of the to-be-measured off-set decoction is compared with the standard fingerprint spectrum obtained by the method. Compared with the traditional HPLC method, the UPLC in the application greatly shortens the online analysis time, saves the material cost and the time cost, and has simple preparation method of the sample to be tested and easy realization of chromatographic conditions.
In summary, the present application has the following beneficial effects:
1. according to the method, the mobile phase composition, the gradient elution program, the flow rate, the detection wavelength and the pretreatment of the sample solution are used for preparing the fingerprint, a traditional Chinese medicine chromatographic fingerprint similarity evaluation system is used for carrying out UPLC fingerprint similarity analysis on 30 batches of scattered soup, the similarity is above 0.90, the method for obtaining the scattered soup UPLC fingerprint is reliable, the operation is simple and convenient, the quality of the scattered soup can be scientifically and effectively evaluated, and the requirements of enterprises on quality control of scattered soup products are met;
2. according to the method for detecting the UPLC fingerprint of the obtained scattered soup, paeoniflorin is used as a reference peak, UPLC fingerprint of 30 batches of scattered soup is established, 13 common peaks are obtained, analysis and identification are carried out on the obtained UPLC common peaks, and components such as sinapine, ferulic acid, paeoniflorin, albiflorin and glycyrrhizic acid are identified, so that the chemical substance basis of the scattered soup is further elucidated.
3. The UPLC fingerprint spectrum method can be used as a control method for controlling the quality of the off-set soup, can be applied to the authenticity identification and component detection of the off-set soup, and can be used for comparing the fingerprint spectrum of the off-set soup to be detected with the standard fingerprint spectrum obtained by the method; compared with the traditional HPLC method, the UPLC in the application greatly shortens the online analysis time, saves the material cost and the time cost, and has simple preparation method of the sample to be tested and easy realization of chromatographic conditions.
Drawings
FIG. 1 is a fingerprint of a control of the present application;
FIG. 2 is a UPLC fingerprint of 30 batches of the off-set soup of the present application;
fig. 3 is a standard fingerprint of the present application.
Detailed Description
The present application is further described in detail below with reference to the attached drawings and examples, and is specifically described as follows: the following examples, in which no specific conditions are noted, are conducted under conventional conditions or conditions recommended by the manufacturer, and the raw materials used in the following examples are commercially available from ordinary sources except for the specific descriptions.
The establishing method of the UPLC fingerprint and the obtained standard fingerprint can be used for detecting a scattered soup preparation, the scattered soup preparation can be a granular preparation, or can be scattered soup freeze-dried powder or a preparation product in the form of an extracting solution, a concentrated solution and other intermediate products in the preparation process of granules after the scattered soup raw materials are mixed, and the scattered soup granular preparation is taken as an example for illustration in the following embodiment.
Example 1
The establishment of the method for measuring the UPLC fingerprint of the scattered decoction comprises the following steps:
1. the preparation of the powder bias soup granule preparation comprises the following steps:
weighing 777g of white peony root, 1554g of szechuan lovage rhizome, 155g of bunge cherry seed, 155g of Chinese thorowax root, 466g of white mustard seed, 311g of nutgrass galingale rhizome, 155g of liquoric root and 78g of dahurian angelica root, adding 8 times of water of the total mass, soaking for 30min, boiling and then decocting for 30min, filtering, adding 6 times of water into filter residues, boiling and then decocting for 25min, filtering, combining the two decoctions, concentrating under reduced pressure at 65 ℃ until the relative density is 1.05-1.10, adding 146g of dextrin, spray-drying to obtain dry paste powder, adding the dextrin to 1000g, uniformly mixing, granulating to 1000g, and obtaining the powder preparation of the polarized soup granule.
The raw materials of different producing areas are randomly taken and respectively prepared into the granule preparation of the scattered soup according to the method, and the number of the granule preparation is respectively SPT212473, SPT212478, SPT212483, SPT212488, SPT212493, SPT212498, SPT212503, SPT212508, SPT212513 and SPT212518 is equal to 30.
2. Establishment of UPLC fingerprint spectrum measuring method of scattered decoction
(a) Preparation of control solution: taking paeoniflorin reference 7mg, adding 10% methanol to prepare a solution containing 0.6mg per 1ml, and taking the solution as reference solution;
(b) Preparation of test solution: taking 1g of a powder bias decoction particle preparation, adding 25ml of 70% methanol, weighing, and performing ultrasonic treatment for 30min, wherein the ultrasonic treatment parameters are as follows: the power is 320w, and the frequency is 40kHz; taking out, cooling, weighing, supplementing the weight of the filtrate with 70% methanol, mixing, and filtering to obtain filtrate as sample solution;
(c) And (3) measuring: 2 μl of each of the reference solution and the sample solution is precisely sucked, and is injected into a liquid chromatograph to collect chromatograms to obtain ultra-high performance liquid chromatograms of the reference solution and 30 batches of the sample solution of the off-set soup, respectively shown in fig. 1 and 2;
in step (c), the liquid chromatography conditions are: octadecylsilane chemically bonded silica is used as a filler, acetonitrile is used as a mobile phase A, and a 0.1% phosphoric acid solution is used as a mobile phase B; the gradient elution procedure was: 0-8min, A2→8%;8-15min, A8%;15-25min, A8-15%; 25-27min, A15-20%; 27-37min, A20→25%;37-60min, A25-70%; the detection wavelength is 230nm, the column temperature is 30 ℃, and the flow rate is 0.3ml/min.
3. Analysis of the atlas
Analyzing fingerprint information of 30 batches of the test sample of the scattered soup to obtain 13 common peaks, wherein the common peaks form fingerprint characteristics of the scattered soup, establishing a UPLC standard fingerprint of the scattered soup as shown in figure 3, calculating relative retention time and relative peak area of each characteristic peak according to retention time and peak area of paeoniflorin as shown in the following tables 1 and 2, and introducing data into a traditional Chinese medicine chromatographic fingerprint similarity evaluation system (2004A version) software evaluation fingerprint similarity graph as shown in the table 3 for fingerprint methodology investigation.
Table 1: relative retention time of each common peak
Continuing with table 1:
table 2: relative peak area of each common peak
Continuing with table 2:
table 3:30 batches of off-set soup and similarity of fingerprint patterns of the control images
As shown in Table 3, the fingerprint of the different batches of the off-set soup is compared with the reference fingerprint, and the similarity is above 0.9.
In conclusion, the fingerprint method established by the application is stable and reliable, can be used as a control method for controlling the quality of the off-set soup, and can be applied to the identification of the authenticity of the off-set soup and the detection of components. When the method is applied, the fingerprint spectrum of the to-be-measured off-set decoction is compared with the standard fingerprint spectrum obtained by the method. Compared with the traditional HPLC method, the UPLC has the advantages that the detection process time is 1h, compared with the detection time of 2h of the traditional HPLC, the on-line analysis time is greatly shortened, the material cost and the time cost are saved, the preparation method of the sample is simple, and the chromatographic condition is easy to realize.
13 common peaks in the fingerprint spectrum of the scattered soup are analyzed, wherein the peak No. 2 is sinapine, the peak No. 3 is albiflorin, the peak No. 4 is paeoniflorin as a reference peak, the peak No. 5 is ferulic acid, the peak No. 7 is glycyrrhizin, the peak No. 8 is isoliquiritigenin, the peak No. 11 is benzoylpaeoniflorin, and the peak No. 12 is glycyrrhizic acid.
4. Precision, repeatability and stability assays
4.1 precision test
The same sample was sampled 6 times in succession, and the relative retention time and relative peak area of each common peak are shown in tables 4 and 5, respectively:
table 4: precision test-relative retention time
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Continuing table 4:
relative retention time Peak 8 Peak 9 Peak No. 10 Peak No. 11 Peak No. 12 Peak No. 13
First needle 1.313 1.372 1.631 1.903 2.267 2.358
Second needle 1.292 1.352 1.613 1.891 2.262 2.344
Third needle 1.314 1.367 1.634 1.905 2.278 2.358
Fourth needle 1.314 1.372 1.631 1.904 2.269 2.356
Fifth needle 1.293 1.354 1.611 1.875 2.241 2.330
Sixth needle 1.309 1.367 1.625 1.891 2.266 2.348
Average value of 1.306 1.364 1.624 1.895 2.264 2.349
Table 5: precision test-relative peak area
Relative retention time Peak 1 Peak No. 2 Peak 3 Peak No. 4 Peak 5 Peak 6 Peak No. 7
First needle 0.171 0.342 0.145 1.000 0.387 0.146 0.148
Second needle 0.172 0.342 0.149 1.000 0.389 0.147 0.152
Third needle 0.169 0.337 0.144 1.000 0.385 0.141 0.146
Fourth needle 0.170 0.353 0.144 1.000 0.382 0.148 0.147
Fifth needle 0.172 0.353 0.144 1.000 0.392 0.149 0.161
Sixth needle 0.174 0.356 0.150 1.000 0.393 0.149 0.149
Average value of 0.171 0.347 0.146 1.000 0.388 0.147 0.151
Continuing with table 5:
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the result shows that the precision of the method is good, and the method meets the fingerprint measurement requirement.
4.2 repeatability test
Samples of the same lot were taken and 6 test sample solutions were prepared in parallel, and the relative retention time and relative peak area of each common peak are shown in tables 6 and 7, respectively:
table 6: repeatability test-relative retention time
Relative retention time Peak 1 Peak No. 2 Peak 3 Peak No. 4 Peak 5 Peak 6 Peak No. 7
Sample 1 0.305 0.735 0.841 1.000 1.098 1.190 1.293
Sample 2 0.303 0.732 0.842 1.000 1.102 1.204 1.287
Sample 3 0.305 0.729 0.838 1.000 1.104 1.203 1.297
Sample 4 0.307 0.732 0.839 1.000 1.107 1.209 1.301
Sample 5 0.305 0.726 0.834 1.000 1.109 1.210 1.302
Sample 6 0.307 0.726 0.835 1.000 1.108 1.210 1.304
Average value of 0.305 0.730 0.838 1.000 1.105 1.204 1.297
Continuing with table 6:
relative retention time Peak 8 Peak 9 Peak No. 10 Peak No. 11 Peak No. 12 Peak No. 13
Sample 1 1.313 1.372 1.631 1.903 2.267 2.358
Sample 2 1.325 1.394 1.650 1.948 2.312 2.412
Sample 3 1.335 1.396 1.667 1.951 2.315 2.411
Sample 4 1.339 1.400 1.680 1.958 2.334 2.419
Sample 5 1.341 1.402 1.677 1.954 2.331 2.415
Sample 6 1.342 1.404 1.677 1.949 2.330 2.413
Average value of 1.332 1.395 1.664 1.944 2.315 2.405
Table 7: repeatability test-relative peak area
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Continuing with Table 7:
relative retention time Peak 8 Peak 9 Peak No. 10 Peak No. 11 Peak No. 12 Peak No. 13
Sample 1 0.071 0.043 0.058 0.049 0.036 0.054
Sample 2 0.068 0.056 0.058 0.048 0.038 0.053
Sample 3 0.070 0.055 0.062 0.051 0.041 0.053
Sample 4 0.071 0.065 0.057 0.049 0.040 0.051
Sample 5 0.071 0.062 0.058 0.049 0.040 0.052
Sample 6 0.070 0.059 0.059 0.050 0.040 0.053
Average value of 0.070 0.057 0.059 0.049 0.039 0.053
The results show that the method is good in repeatability.
4.3 stability test
Taking the same sample, and injecting the sample for 0,3,6,9,12,15h,18h and 24h respectively, wherein the relative retention time and the relative peak area of each common peak are shown in tables 8 and 9 respectively:
table 8: stability test-relative retention time
Continuing with Table 8:
relative retention time Peak 8 Peak 9 Peak No. 10 Peak No. 11 Peak No. 12 Peak No. 13
0h 1.313 1.372 1.631 1.903 2.267 2.358
3h 1.314 1.373 1.634 1.905 2.278 2.358
6h 1.313 1.374 1.611 1.875 2.281 2.330
9h 1.317 1.376 1.637 1.893 2.273 2.354
12h 1.336 1.397 1.671 1.956 2.321 2.405
15h 1.341 1.403 1.673 1.950 2.321 2.405
18h 1.326 1.386 1.661 1.938 2.316 2.400
24h 1.325 1.384 1.645 1.920 2.282 2.365
Average value of 1.325 1.385 1.647 1.920 2.296 2.374
Table 9: stability test-relative peak area
Relative retention time Peak 1 Peak No. 2 Peak 3 Peak No. 4 Peak 5 Peak 6 Peak No. 7
0h 0.171 0.342 0.145 1.000 0.387 0.146 0.148
3h 0.169 0.337 0.144 1.000 0.385 0.141 0.146
6h 0.172 0.353 0.144 1.000 0.392 0.149 0.161
9h 0.173 0.341 0.151 1.000 0.391 0.147 0.149
12h 0.171 0.338 0.161 1.000 0.384 0.147 0.148
15h 0.172 0.338 0.164 1.000 0.385 0.147 0.147
18h 0.172 0.341 0.162 1.000 0.390 0.147 0.152
24h 0.190 0.376 0.164 1.000 0.433 0.162 0.165
Average value of 0.174 0.346 0.156 1.000 0.394 0.149 0.153
Continuing table 9:
the results indicated that the test solution was stable over 24 hours.
The present embodiment is merely illustrative of the present application and is not intended to be limiting, and those skilled in the art, after having read the present specification, may make modifications to the present embodiment without creative contribution as required, but is protected by patent laws within the scope of the claims of the present application.

Claims (5)

1. A method for detecting a UPLC fingerprint of a scattered soup is characterized by comprising the following steps:
(a) Preparation of control solution: taking paeoniflorin reference substance, adding methanol solution to prepare reference substance solution;
(b) Preparation of test solution: adding methanol solution into the decoction, weighing, ultrasonic treating for 30min, taking out, cooling, weighing again, supplementing the reduced weight with methanol solution, mixing, and filtering to obtain filtrate as sample solution;
(c) And (3) measuring: precisely sucking the reference solution and the sample solution respectively, injecting into an ultra-high performance liquid chromatograph, and collecting chromatograms;
(d) Introducing the integral signals of the fingerprints of the sample solution of the scattered soup obtained in the step (c) into evaluation software, and selecting chromatographic peaks existing in different batches of scattered chromatograms as common peaks to obtain standard fingerprints of the scattered soup;
the preparation of the depolarizing soup in the step (b) is prepared by the following method:
weighing 777g of white peony root, 1554g of szechuan lovage rhizome, 155g of bunge cherry seed, 155g of Chinese thorowax root, 466g of white mustard seed, 311g of nutgrass galingale rhizome, 155g of liquoric root and 78g of dahurian angelica root, adding 8 times of water of the total mass, soaking for 30min, boiling, decocting, filtering, adding 6 times of water into filter residues, boiling, decocting, filtering, combining the two decoctions, concentrating under reduced pressure at 65 ℃ until the relative density is 1.05-1.10, adding dextrin, spray-drying into dry paste powder, adding dextrin, uniformly mixing, granulating to 1000g, and obtaining a biased soup granule preparation;
wherein in step (c), the liquid chromatography conditions are: octadecylsilane chemically bonded silica is used as a filler, acetonitrile is used as a mobile phase A, and phosphoric acid solution is used as a mobile phase B; gradient elution: 0-8min, A2→8%;8-15min, A8%;15-25min, A8-15%; 25-27min, A15-20%; 27-37min, A20→25%;37-60min, A25-70%; the detection wavelength is 230nm;
13 common peaks, namely sinapine in peak No. 2, paeoniflorin in peak No. 3, paeoniflorin in peak No. 4, ferulic acid in peak No. 5, glycyrrhizin in peak No. 7, isoliquiritigenin in peak No. 8, benzoylpaeoniflorin in peak No. 11 and glycyrrhizic acid in peak No. 12;
in the standard fingerprint, paeoniflorin is used as a reference peak, and the relative retention time of the obtained common peak is as follows: peak 1:0.280; peak 2:0.797; peak 3:0.897; peak 4:1.000; peak 5:1.055; peak 6:1.110; peak 7:1.211; peak 8:1.231; peak 9:1.262; peak 10:1.510; peak 11:1.822; peak 12:2.038; peak 13:2.126; or (b)
In the standard fingerprint, paeoniflorin is used as a reference peak, and the relative peak area of the obtained common peak is as follows: peak 1:0.222; peak 2:0.442; peak 3:0.162; peak 4:1.000; peak 5:0.347; peak 6:0.208; peak 7:0.109; peak 8:0.064; peak 9:0.042; peak 10:0.041; peak 11:0.060; peak 12:0.036; peak 13:0.046.
2. the method for detecting the UPLC fingerprint of the scattered soup according to claim 1, which is characterized by comprising the following steps: in step (b), the sonication parameters are: the power is 310-330w, and the frequency is 35-45kHz.
3. The method for detecting the UPLC fingerprint of the scattered soup according to claim 1, which is characterized by comprising the following steps: in the step (d), the similarity between the fingerprint of the sample and the fingerprint of the reference sample is not lower than 0.90.
4. A standard fingerprint spectrum of a scattered soup UPLC is characterized in that: a method according to any one of claims 1-3.
5. The use of standard fingerprint of the yao-decoction UPLC in the detection of yao-decoction preparation as claimed in claim 4.
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