CN109374764B - HPLC fingerprint spectrum and main component content determination method for eight-ingredient Longzui granules - Google Patents
HPLC fingerprint spectrum and main component content determination method for eight-ingredient Longzui granules Download PDFInfo
- Publication number
- CN109374764B CN109374764B CN201811226190.3A CN201811226190A CN109374764B CN 109374764 B CN109374764 B CN 109374764B CN 201811226190 A CN201811226190 A CN 201811226190A CN 109374764 B CN109374764 B CN 109374764B
- Authority
- CN
- China
- Prior art keywords
- ingredient
- sinomenine
- granules
- concentration
- solution
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention provides a method for qualitative and quantitative analysis of HPLC (high Performance liquid chromatography) fingerprint spectrums of main components of eight-flavor Longzun particles. The method adopts 70% methanol ultrasonic-assisted extraction, uses octadecylsilane chemically bonded silica as a stationary phase and acetonitrile-0.1% phosphoric acid aqueous solution as a mobile phase, and can simultaneously determine the characteristic qualitative fingerprint spectrum and the contents of 7 main active ingredients of the preparation under one chromatographic condition. Scientific experiments prove that the method is simple, convenient and quick to operate, good in repeatability and stability, strong in fingerprint characteristic, accurate in content determination result and capable of being used for quality control of eight-ingredient Longzui particle products.
Description
Technical Field
The invention relates to a method for simultaneously carrying out qualitative and quantitative analysis on main components of eight-ingredient Longzui granules, in particular to a method for simultaneously carrying out quantitative detection on the qualitative and 7 main efficacy-related components of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen by HPLC fingerprint chromatography, belonging to the field of analysis and detection of compound traditional Chinese medicine preparations.
Background
The eight-ingredient Longzui granules are Zhuang patent drugs researched and developed according to clinical proved recipes compatible with Zhuang medical theory on the basis of mining and summarizing traditional medicines for treating rheumatoid arthritis and the connotation of Zhuang medical toxicology. The prescription mainly comprises eight medicines of asiatic toddalia root, rhinestone, shinyleaf pricklyash root, caulis sinomenii, caulis spatholobi, Japanese bauhinia stem, Chinese alangium and hispid fig root, has the effects of resisting inflammation, easing pain, resisting rheumatism and immunosuppression, and has obvious curative effect on rheumatoid arthritis. The bauhinia championii, the suberect spatholobus stem and the like in the prescription are commonly used for treating rheumatic diseases, the bauhinia championii has strong analgesic and anti-inflammatory activities and is a special common medicine for Zhuang Yi rheumatism diseases, and the suberect spatholobus stem has the effects of promoting blood circulation, enriching blood, dredging collaterals, dispelling wind and removing dampness; caulis Sinomenii has effects of dispelling pathogenic wind, removing blood stasis, promoting diuresis and relieving pain, and radix fici Simplicissimae can invigorate spleen, invigorate qi, eliminate dampness, relax muscles and tendons, promote qi circulation and relieve pain. Modern pharmacological research shows that the total alkaloids, ethanol extract and water extract of Toddalia asiatica have analgesic and anti-inflammatory effects; radix Zanthoxyli has effects of relieving swelling and pain, and salidroside in caulis Sargentodoxae (radix Dactylicapni) has effects of relieving fatigue, resisting inflammation, resisting aging, and regulating immunity, and is one of active ingredients of radix Dactylicapni; the sinomenine has antiinflammatory, analgesic, and antirheumatic effects; psoralen is often used as a quality control index for Ficus simplicissima lour; the hesperidin has pharmacological activities of resisting inflammation, regulating immunity and the like; the nitidine chloride has obvious effect on anti-inflammation; the gallic acid and protocatechuic acid have obvious effects of resisting oxidation and inhibiting platelet aggregation; catechin has certain effect of promoting hemopoietic cell proliferation, and is the main material basis of caulis Spatholobi for replenishing blood and promoting blood circulation. The gallic acid has antibacterial, antioxidant and antitumor pharmacological effects. The above components are closely related to the efficacy of the preparation. So far, no method for simultaneously measuring the fingerprint spectrum which can embody the characteristic information of the eight dragons particle medicinal materials and simultaneously measuring the content of main characteristic components in the preparation is reported by only using one chromatographic system.
Disclosure of Invention
The invention provides a method for simultaneously determining a characteristic fingerprint spectrum capable of reflecting the information of main medicinal materials of eight-flavor Longjing particles and simultaneously determining the content of 7 main characteristic components related to efficacy in a preparation, wherein 70% methanol is adopted for ultrasonic-assisted extraction, octadecylsilane chemically bonded silica is taken as a stationary phase, acetonitrile-0.1% phosphoric acid aqueous solution is taken as a mobile phase, and the characteristic qualitative fingerprint spectrum and the content of 7 main active components of the preparation can be simultaneously determined by only one chromatographic condition. Scientific experiments prove that the method is simple, convenient and quick to operate, good in repeatability and stability, strong in fingerprint characteristic, accurate in content determination result and capable of being used for quality control of eight-ingredient Longzui particle products.
The detection method for detecting the characteristic fingerprint spectrum and the content of 7 efficacy-related components of the eight-ingredient Longzui granules comprises the following steps:
1. preparation of control solutions: precisely weighing appropriate amount of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen reference substances, dissolving with methanol, and respectively making into reference substance stock solutions with appropriate concentrations. And putting a proper amount of the solution into a same measuring flask, adding methanol for dilution, preparing a mixed reference substance solution containing gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen with certain concentrations, and storing in a refrigerator for later use.
2. Preparation of a test solution: precisely weighing an appropriate amount of the eight-ingredient Longzu granules, placing the eight-ingredient Longzu granules in a conical flask with a plug, precisely adding an appropriate amount of 70% methanol, weighing, ultrasonically treating, cooling to room temperature, weighing again, supplementing the lost weight with 70% methanol, shaking up, filtering, centrifuging the filtrate, and taking the supernatant as a test solution.
3. Chromatographic conditions are as follows: performing gradient elution by using octadecylsilane chemically bonded silica as a stationary phase and acetonitrile-0.1% phosphoric acid aqueous solution as a mobile phase; the column temperature is 25-30 ℃; the flow rate is 1 ml/min; the detector is an ultraviolet detector, and the detection wavelengths are 270nm and 245nm respectively.
4. The determination method comprises the following steps: accurately sucking appropriate amount of mixed reference solution and test solution of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen respectively, injecting into high performance liquid chromatograph according to the above chromatographic conditions, measuring corresponding peak area, and calculating to obtain the final product.
Through research, the optimized detection method for detecting the characteristic fingerprint spectrum and the content of 7 efficacy-related components of the eight-ingredient Longzui granules is implemented according to the following steps:
1. preparation of control solutions: precisely weighing appropriate amount of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen reference substances, dissolving with methanol, and respectively making into reference substance stock solutions with appropriate concentrations. And putting a proper amount of the solution into the same measuring flask, adding methanol for dilution to prepare a mixed reference solution containing 0.05-0.10 mg/mL of gallic acid, 0.005-0.01 mg/mL of protocatechuic acid and psoralen, 0.1-0.3 mg/mL of sinomenine, salidroside and catechin and 0.01-0.05 mg/mL of hesperidin, and storing the mixed reference solution in a refrigerator for later use.
2. Preparation of a test solution: precisely weighing 3g of Bawei Longzu granules, placing the granules in a conical flask with a plug, precisely adding 50mL of 70% methanol, weighing, carrying out ultrasonic treatment (power 200W and frequency 40kHz) for 20min, cooling, weighing again, supplementing the lost weight with 70% methanol, shaking up, filtering, centrifuging the subsequent filtrate, and taking the supernatant to obtain a test solution;
3. chromatographic conditions and chromatographic applicability test: the method comprises the following steps of taking a Phenomenex Gemini C18(250mm multiplied by 4.6mm i.d., 5 mu m) chromatographic column as a stationary phase and acetonitrile (A) -0.1% phosphoric acid water (B) as a mobile phase, and carrying out gradient elution according to volume percentage conditions: 0-10 min, 6% A; 10-20 min, 12% A; 20-30 min, 19% A; 30-40 min, 27% A; 40-50 min, 35% A; 45% of A for 50-60 min; the column temperature is 25-30 ℃; the flow rate is 1 ml/min; the detector is an ultraviolet detector, and the detection wavelength is 0-50 min: 270nm, 50-60 min: 245 nm; the sample volume is 10 mu L; the column efficiency of the chromatographic column is not less than 30000 calculated by sinomenine.
4. The determination method comprises the following steps: respectively and precisely sucking 10 μ L of the mixed reference solution and the sample solution containing gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen at certain concentrations, injecting into a high performance liquid chromatograph under the above chromatographic conditions, measuring corresponding peak area, and calculating to obtain the final product.
Compared with the prior art, the invention has the main advantages and positive effects that:
(1) the detection method of the invention adopts the wavelength conversion technology, and can simultaneously determine the characteristic fingerprint spectrum of the preparation and the contents of 7 efficacy-related components of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen in the preparation by only using one chromatographic condition.
(2) The detection method of the invention is simple, convenient and quick, is easy to operate, and has no fussy operation process.
(3) The detection method has the characteristics of simplicity, convenience, rapidness, stability, good repeatability and reproducibility, high accuracy and easiness in mastering and operating.
The detection method of the present invention is a feasible method obtained through a large number of experiments, and has repeatability and better stability, and other embodiments of the present invention will be clear to those skilled in the art according to the technical content disclosed by the present invention, but the detection method of the present invention is within the protection scope of the present invention.
Description of the drawings:
FIG. 1 HPLC-UV chromatogram of 7 mixed reference solutions such as sinomenine
FIG. 2 shows HPLC-UV spectrum for measuring content of 7 components in test solution of eight-ingredient Longzui granule
FIG. 3 is an overlay of HPLC finger print of eight ingredients of Longzu granules
FIG. 4 shows the common HPLC fingerprint pattern of the eight ingredient Longzui granules
The specific implementation mode is as follows:
the present invention will be further described with reference to the following examples, which are intended to be illustrative only and are not to be construed as limiting the invention.
Example determination of the content of 7 major ingredients in an eight ingredient Longzui granule
1 Instrument and reagent
1.1 Instrument: agilent-1260 high performance liquid chromatograph; a diode array ultraviolet detector; ultrasonic cleaners (KQ5200, ultrasonic instruments ltd, kunshan); centrifuge (TGL-16G, Shanghai' an pavilion scientific instruments factory); electronic balances (one hundred thousand, SQP, sydows scientific instruments (beijing) ltd); a Misibo ultra-pure water machine and the like.
1.2 reagent: a \ "control product Gallic acid (Gallic acid) (110831-201605, content 90.8%), Protocatechuic acid (protocatholic acid) (110809-201205, content 99.9%), Sinomenine (Sinomenine) (110774-200507, content 99.1%), Salidroside (110818-2015007, content 99.4%), Catechin ((+) -Catechin) (110877-201604, content 99.2%), Hesperidin (Hesperidin) (110721-201617, content 96.7%) and Psoralen (Psoralen) (1109-7317, content 99.1%) were purchased from China food and drug institute; acetonitrile (chromatically pure; Fisher company); methanol, phosphoric acid (analytical grade, chemical reagents of national drug group, ltd.); the water is ultrapure water. 10 batches of eight-ingredient Longzui granules (10 g/bag, each bag is equivalent to 35 g of raw medicinal materials) are provided by pharmaceutical factories of Guangxi Chinese medicinal university; the experimental raw medicinal materials of the Chinese alangium root, the rhubard, the asiatic toddalia root, the suberect spatholobus stem, the shinyleaf pricklyash root, the kakko yushun, the five-finger wild peach and the caulis sinomenii are purchased from auspicious Chinese medicinal decoction piece Limited liability company in Guangxi Yulin city, and are identified by the assistant principal and subordinate pharmacist of Rouyou in the pharmaceutical factory quality assurance department of Guangxi Chinese medicinal university, and the 8 medicinal materials of the asiatic toddalia root, the rhubard, the pricklyash root and the like all meet the relevant requirements of the quality standard of Guangx.
2 methods and results
2.1 chromatographic conditions and System suitability test
The method comprises the following steps of taking a Phenomenex Gemini C18(250mm multiplied by 4.6mm i.d., 5 mu m) chromatographic column as a stationary phase and acetonitrile (A) -0.1% phosphoric acid water (B) as a mobile phase, and carrying out gradient elution according to volume percentage conditions: 0-10 min, 6% A; 10-20 min, 12% A; 20-30 min, 19% A; 30-40 min, 27% A; 40-50 min, 35% A; 45% of A for 50-60 min; the column temperature is 25-30 ℃; the flow rate is 1 ml/min; the detector is an ultraviolet detector, and the detection wavelength is 0-50 min: 270nm, 50-60 min: 245 nm; the sample volume is 10 mu L; the column efficiency of the chromatographic column is not less than 30000 calculated by sinomenine. Under this chromatographic condition, the other components of the recipe were not interfering with the assay. HPLC-UV spectrums of mixed reference substance solution and sample solution of eight ingredient LONGJING granule containing 7 main ingredients are respectively shown in figure 1 and figure 2.
2.2 preparation of reference and test solutions
2.2.1 preparation of Mixed control solutions: precisely weighing appropriate amount of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen as reference, and dissolving in methanol to obtain reference stock solution. And putting a proper amount of the solution into a same measuring flask, adding methanol for dilution to prepare a mixed reference substance solution containing gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen with the concentrations of 0.0696mg/mL, 0.0087mg/mL, 0.2046mg/mL, 0.2564mg/mL, 0.1004mg/mL, 0.0243mg/mL and 0.0071mg/mL respectively, and storing the mixed reference substance solution in a refrigerator for later use.
2.2.2 preparation of test solutions: precisely weighing 3g of eight-ingredient Longzui granules, placing the eight-ingredient Longzui granules in a conical flask with a plug, precisely adding 50mL of 70% methanol, weighing, ultrasonically treating for 20min (power 200W and frequency 40kHz), cooling to room temperature, weighing again, supplementing the lost weight with 70% methanol, shaking up, filtering, centrifuging the subsequent filtrate, and taking the supernatant as a test solution.
2.3 assay: respectively and precisely sucking 10 μ l of the mixed reference solution and the test solution containing gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen at certain concentrations, injecting into a high performance liquid chromatograph under the above chromatographic conditions, measuring corresponding peak areas, and calculating to obtain the final product.
2.4 methodological considerations
2.4.1 System suitability test: respectively taking a reference substance solution and a sample solution, carrying out sample injection analysis under the chromatographic condition of '2.1', and detecting by using a diode array detector, wherein the results show that the chromatographic peak retention time of 7 components of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen is consistent with the retention time of the UV spectrum and the corresponding chromatographic peak retention time in the sample and the UV spectrum, the separation degree of each peak is good, the tailing factors are all between 0.95 and 1.05, the determination of the 7 components by other components in the sample is not interfered, the theoretical plate number of a chromatographic column is not less than 30000 calculated by sinomenine, and the details are shown in an attached figure 1 and an attached figure 2.
2.4.2 investigation of the Linear relationship: taking the mixed reference substance solution under the item of '2.2.1', respectively and precisely taking 4, 6, 8, 10, 12 and 14 mu L of the mixed reference substance solution by using an automatic sample injector, injecting the solution into a chromatograph for analysis, carrying out sample injection measurement according to the chromatographic condition under the item of '2.1', recording the peak areas of corresponding chromatographic peaks, respectively taking the peak areas Y as vertical coordinates and the sample injection amount (mu g) X as horizontal coordinates, and solving a linear regression equation and a linear correlation coefficient (r), wherein the results are shown in a table 1-1.
TABLE 1-1 regression equation and Linear Range
2.4.2 precision test: taking the mixed reference substance solution under the item of 2.2.1, measuring according to the chromatographic condition of 2.1, continuously injecting samples for 6 times, respectively measuring the RSD of the peak areas of the gallic acid, the protocatechuic acid, the sinomenine, the salidroside, the catechin, the hesperidin and the psoralen to be 1.3%, 0.77%, 0.87%, 1.1%, 0.94% and 1.2%, and displaying that the precision of the instrument is good.
2.4.3 stability test: taking eight-ingredient Longzui granules (batch No. 20170501), preparing a sample solution according to the method under the item '2.2.2', measuring according to the '2.1' chromatographic condition, and respectively measuring the RSD of the peak areas of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen at 0h, 2h, 4h, 8h, 12h and 24h in sequence, wherein the RSD respectively shows that the sample solution is stable within 24h, and the RSD respectively shows that the peak areas of the gallic acid, the protocatechuic acid, the sinomenine, the rhodioside, the catechin, the hesperidin and the psoralen are 1.2%, 1.1.1%, 1.4%, 1..
2.4.4 repeatability tests: precisely weighing 6 parts of eight-ingredient Longjing granules (batch No. 20170501), 3.0g each, preparing 6 parts of test solution by the method under item '2.2.2', and measuring according to the chromatographic condition under item '2.1', wherein the average contents of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen peaks are respectively as follows: 0.6959mg/g, 0.1364mg/g, 3.5620mg/g, 3.6991mg/g, 1.2136mg/g, 0.3234mg/g and 0.07650mg/g, and RSD is 0.71%, 1.9%, 2.0%, 1.5% and 1.5%, respectively, which shows that the method has good repeatability.
2.4.5 sample recovery test: taking about 1.5g of Bawei Longjing granules (batch No. 20170501) (the average contents of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen are 0.6959, 0.1364, 3.5620, 3.6991, 1.2136, 0.3234 and 0.07650mg/g respectively), precisely weighing 6 parts in total, and precisely adding 1.0500mg, 0.2050mg, 5.5000mg, 5.6000mg, 1.8200mg, 0.4800mg and 0.1200 mg of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen reference substances respectively. The sample solution was prepared according to the method under item "2.2.2", and the sample recovery rates of the respective components were calculated as 103.6%, 102.1%, 102.5%, 99.6%, 103.2%, 99.2%, and 98.5%, and RSD as 1.1%, 1.0%, 1.4%, 1.7%, 1.4%, 0.92%, and 1.3%, respectively, as measured under the chromatographic conditions under item "2.1", and the respective peak areas were measured, indicating that the accuracy of the method was good, see tables 1-2.
Table 1-27 component recovery test results (n ═ 6)
2.5 sample determination: taking 2 parts of each of 10 batches of eight-ingredient Longzui granules, each of which is about 3g, precisely weighing, preparing a sample solution according to the method under the item 2.2.2, determining according to the chromatographic condition under the item 2.1, and calculating the average content of each component in the sample according to the peak area, wherein the result is shown in tables 1-3.
Table 1-3 results of content measurement of sample (n ═ 2) (mg/g)
Example HPLC fingerprinting study and determination of twenty-eight ingredient Longzui granules
1 instrument and reagent: the same as the first embodiment.
2, method and result: the same as the first embodiment.
2.1 chromatographic conditions and System suitability test: the same as the first embodiment.
2.2 preparation of reference substance and test solution: the same as the first embodiment.
2.3 assay: the same as the first embodiment.
2.4 HPLC fingerprint methodology investigation and determination of eight ingredient Longzui granules
2.4.1 stability test: taking eight-flavor Longjing particles (lot number 20170501), preparing a test solution according to the method under the item '2.2.2', under the chromatographic condition of the item '2.1', respectively measuring at 0, 2, 4, 8, 12 and 24h, and taking the retention time of the No. 6 peak (sinomenine) and the chromatographic peak area as references, calculating to obtain that the relative retention time RSD of the other 11 common peaks except the reference peak sinomenine is between 0.33 and 1.8 percent, and the relative peak area RSD is between 0.88 and 1.7 percent, wherein the experimental result shows that the eight-flavor Longjing particle test solution is stable within 24h and meets the related requirements and regulations of fingerprint determination.
2.4.2 precision test: taking eight-ingredient Longjing particles (batch number 20170501), preparing a test solution according to the method under the item '2.2.2', continuously sampling for 6 times under the chromatographic condition of the item '2.1', determining an HPLC chromatogram, and calculating to obtain the relative retention time RSD of 11 common peaks except for sinomenine, wherein the relative retention time RSD is between 0.40 and 1.6 percent and the relative peak area RSD is between 0.62 and 1.8 percent by taking the retention time and the chromatographic peak area of the 6 peak (sinomenine) as references, thereby indicating that the precision of the instrument is good and meeting the related requirements and regulations of fingerprint determination.
2.4.3 repeatability tests: taking 6 parts of eight-ingredient Longjing particle samples (lot 20170501) of the same batch, each of which is about 3g, precisely weighing, preparing according to a method under the item '2.2.2', measuring under the chromatographic condition of the item '2.1', and calculating to obtain the relative retention time RSD of the relative retention time of the other 11 common peaks except for sinomenine to be between 0.18 and 1.2 percent and the relative peak area RSD to be between 0.33 and 1.8 percent by taking the retention time of the peak (sinomenine) 6 and the chromatographic peak area as reference, wherein the results show that the method has good repeatability and meets the requirement of fingerprint spectrum.
2.4.4 establishment of HPLC fingerprint of eight ingredient Longzui granules
Taking 10 batches of eight dragons of particle samples, preparing a test solution according to the method under the item 2.2.2, and measuring and analyzing under the chromatographic condition under the item 2.1, as shown in the attached figure 3. Establishing common mode chromatogram (average value) of HPLC finger prints of the eight-ingredient Longzui granule, and analyzing by using a Chinese medicinal chromatogram finger print similarity evaluation system, as shown in figure 4. 12 common peaks are formed on the contrast map, wherein the No. 6 chromatographic peak (sinomenine) in the contrast map has better separation degree, higher chromatographic response value and intermediate retention time, so the No. 6 peak (sinomenine) is selected as the reference peak. The similarity evaluation of the fingerprints of the eight-ingredient Longzui granules adopts a traditional Chinese medicine chromatogram fingerprint similarity evaluation system (2012.130723 edition), and the similarity calculation results of 10 batches of the eight-ingredient Longzui granules are shown in table 2-1 after the detection data of the fingerprints of the eight-ingredient Longzui granules are processed and analyzed. As can be seen from Table 2-1, the similarity of the eight ingredient Longzui granules in each batch was 0.989 or more, and the similarity was good. The consistency of the index components contained in each batch of samples is better.
TABLE 2-110 similarity of eight ingredient Longzui granules to control finger print
2.4.5 analysis of assignment of main common characteristic peaks of HPLC fingerprint
Precisely absorbing the reference substance solutions of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen respectively, analyzing and measuring according to the chromatographic condition under the item '2.1', simultaneously recording chromatogram and ultraviolet absorption spectrum of each peak by using a diode array detector, comparing with each main common peak in the fingerprint common mode, and displaying that the retention time and spectrum of characteristic peaks 3, 5, 6, 7, 9, 10 and 12 in the fingerprint common mode are respectively consistent with the retention time and ultraviolet absorption spectrum of gallic acid, protocatechuic acid, sinomenine, rhodioside, catechin, hesperidin and psoralen reference substances, thereby confirming that the peak 3 of the granule chromatogram of the eight-flavor Chinese Longzuki is gallic acid, the peak 5 is protocatechuic acid, the peak 6 is sinomenine, the peak 7 is rhodioside and the peak catechin 9, peak 10 is hesperidin and Peak 12 is psoralen.
Claims (2)
1. A can be used for eight ingredient longzhi zuo drilling granulometric principal ingredients qualitative and quantitative simultaneous detection method, said method can determine eight ingredient longzhi zuoji drilling granulometric HPLC fingerprint and eight ingredient longzhi zuoji drilling granulometric content of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen 7 kinds of principal ingredients at the same time only with a chromatographic condition, characterized by that the detection method includes the following steps:
(1) chromatographic conditions are as follows: by Phenomenex Gemini C18The column is a fixed phase, and the specification of the column is 250mm multiplied by 4.6mm i.d., 5 mu m; taking acetonitrile and 0.1% phosphoric acid aqueous solution as mobile phases, wherein the acetonitrile is A phase, the 0.1% phosphoric acid aqueous solution is B phase, and carrying out gradient elution, wherein the volume percentage conditions of the gradient elution are as follows: 0-10 min, 6% A; 10-20 min, 12% A; 20-30 min, 19% A; 30-40 min, 27% A; 40-50 min, 35% A; 45% of A for 50-60 min; the column temperature is 25-30 ℃; the flow rate is 1 ml/min; the detector is an ultraviolet detector, and the detection wavelength is 0-50 min: 270nm, 50-60 min: 245 nm; sample size is 10 muL; the column efficiency of the chromatographic column is not less than 30000 calculated by sinomenine;
(2) preparation of control solutions: precisely weighing appropriate amount of gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen reference substances, dissolving with methanol, and respectively making into reference substance stock solutions with appropriate concentration; taking a proper amount of each reference substance stock solution, placing the reference substance stock solution into a same measuring flask, adding methanol for dilution to prepare a mixed reference substance solution with the concentration of gallic acid being 0.05-0.10 mg/mL, the concentration of protocatechuic acid being 0.005-0.01 mg/mL, the concentration of psoralen being 0.005-0.01 mg/mL, the concentration of sinomenine being 0.1-0.3 mg/mL, the concentration of salidroside being 0.1-0.3 mg/mL, the concentration of catechin being 0.1-0.3 mg/mL and the concentration of hesperidin being 0.01-0.05 mg/mL, placing the mixed reference substance solution into a refrigerator for storage and later use;
(3) preparing a test solution of eight-ingredient Longzui granules: precisely weighing 3g of Bawei Longzui granules, placing the granules in a conical flask with a plug, precisely adding 50mL of 70% methanol, weighing, ultrasonically treating the granules for 20min at the power of 200W and the frequency of 40kHz, cooling, weighing again, supplementing the lost weight with 70% methanol, shaking uniformly, and filtering; centrifuging the filtrate, and collecting supernatant to obtain test solution;
(4) measurement method
Respectively and precisely sucking 10 μ L of the mixed reference solution containing gallic acid, protocatechuic acid, sinomenine, salidroside, catechin, hesperidin and psoralen at a certain concentration and the sample solution of the eight-ingredient rhinestone particles, injecting into a high performance liquid chromatograph under the chromatographic conditions in the step (1), measuring the corresponding peak area, and calculating to obtain the target product.
2. The simultaneous detection method for qualitative and quantitative analysis of the main ingredients of eight ingredient rhizoma paridis granule as claimed in claim 1, wherein the eight ingredient rhizoma paridis granule is prepared from eight herbs including radix Toddaliae Asiaticae, radix Dactylicapni, radix Zanthoxyli, caulis Sinomenii, caulis Spatholobi, caulis Bauhihiae Championii, Chinese Alangium and hispid Fig.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811226190.3A CN109374764B (en) | 2018-10-19 | 2018-10-19 | HPLC fingerprint spectrum and main component content determination method for eight-ingredient Longzui granules |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811226190.3A CN109374764B (en) | 2018-10-19 | 2018-10-19 | HPLC fingerprint spectrum and main component content determination method for eight-ingredient Longzui granules |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN109374764A CN109374764A (en) | 2019-02-22 |
| CN109374764B true CN109374764B (en) | 2021-05-14 |
Family
ID=65401122
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201811226190.3A Active CN109374764B (en) | 2018-10-19 | 2018-10-19 | HPLC fingerprint spectrum and main component content determination method for eight-ingredient Longzui granules |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN109374764B (en) |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112816603A (en) * | 2021-01-05 | 2021-05-18 | 天津中医药大学 | Bone-strengthening joint pill fingerprint spectrum and method for detecting 18 components in bone-strengthening joint pill fingerprint spectrum |
| CN112924600B (en) * | 2021-03-23 | 2023-11-03 | 贵州益佰制药股份有限公司 | Fingerprint spectrum establishment method of preparation for treating joint swelling and pain caused by rheumatic arthralgia |
| CN112986458A (en) * | 2021-03-23 | 2021-06-18 | 贵州益佰制药股份有限公司 | Content determination method of Jingu lotus preparation |
| CN114236034B (en) * | 2021-12-15 | 2024-03-29 | 贵州恒霸药业有限责任公司 | Method for detecting asiatic toddalia root blood medicinal material |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| UA78194C2 (en) * | 2000-12-08 | 2007-03-15 | Council Scient Ind Res | Method for revealing and identification of active components in extracts by chromatographic fingerprinting of "fingerprints" and processor of program data processing chromatograms |
| WO2015042852A1 (en) * | 2013-09-27 | 2015-04-02 | 四川川大华西药业股份有限公司 | Method for detecting pulse-activating injection intermediates |
| CN104825619A (en) * | 2014-11-04 | 2015-08-12 | 广西中医药大学 | Medicine composition for treating rheumatoid arthritis and preparation method thereof |
| CN104666700A (en) * | 2015-02-09 | 2015-06-03 | 临沂大学 | Traditional Chinese medicine composition for treating fracture and quality detection method of traditional Chinese medicine composition |
| CN107543886A (en) * | 2017-09-01 | 2018-01-05 | 广西中医药大学制药厂 | A kind of eight tastes dragon bores three kinds of active component content methods for measuring simultaneously in particle |
| CN108645946A (en) * | 2018-05-11 | 2018-10-12 | 中国人民解放军新疆军区药品仪器检验所 | The Quality evaluation method of its particle of lotus prestige arna |
-
2018
- 2018-10-19 CN CN201811226190.3A patent/CN109374764B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN109374764A (en) | 2019-02-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN109374764B (en) | HPLC fingerprint spectrum and main component content determination method for eight-ingredient Longzui granules | |
| CN107153098B (en) | Method for measuring fingerprint spectrum of traditional Chinese medicine composition | |
| CN104849364B (en) | Canzhiling oral solution fingerprint map building method, fingerprint map and application thereof | |
| CN111999423B (en) | Establishment method and application of neck pain granule fingerprint spectrum | |
| CN107860832B (en) | Method for establishing fingerprint of compound rhubarb clear pancreas soup | |
| Wu et al. | Simultaneous quantitative determination of nine active chemical compositions in traditional Chinese medicine Glycyrrhiza by RP-HPLC with full-time five-wavelength fusion method | |
| CN112782308B (en) | Method for establishing HPLC fingerprint of eucommia ulmoides Butiansu pill | |
| CN112666277B (en) | HPLC (high Performance liquid chromatography) characteristic spectrum construction and detection method for rhizoma cyperi medicinal materials, decoction pieces, standard decoction and formula granules | |
| CN103969372A (en) | Content determination and identification method for Xinkeshu capsules | |
| CN104007198B (en) | A kind of glossy ganoderma emperor's preparation HPLC standard finger-print and construction method thereof and application | |
| CN109709222B (en) | Component detection method of Ganmaoling and compound Ganmaoling | |
| CN109115904B (en) | Construction method and application of UPLC fingerprint of Dingkundan | |
| CN109142563B (en) | Method for constructing tortoise age collection UPLC fingerprint and application thereof | |
| CN102068553A (en) | Method for constructing high performance liquid chromatographic (HPLC) fingerprint of Rupixiao preparation | |
| CN105486761A (en) | Method for determining scutelloside content in traditional Chinese medicine granules | |
| CN109781884B (en) | Establishing method of Qianliexin capsule fingerprint and fingerprint thereof | |
| CN103604898A (en) | Fingerprint spectrum detection method and fingerprint spectrum of Yixinshu preparation | |
| CN109856262B (en) | Method for qualitative and quantitative analysis of main components of Erding preparation simultaneously | |
| CN113341007A (en) | Method for measuring contents of multiple components in whole Chinese date seed nerve-soothing capsule based on HPLC (high performance liquid chromatography) characteristic spectrum | |
| CN107894466B (en) | Determination method of HPLC fingerprint of Jinsangqi antitoxic preparation and quality control method of Jinsangqi antitoxic preparation | |
| CN105203469A (en) | Detection method of medicament for treating femoral head necrosis | |
| CN111751472A (en) | Quality evaluation method of sophora and scutellaria ointment | |
| CN112798721B (en) | Fingerprint construction method and application of traditional Chinese medicine preparation for treating cervical spondylosis | |
| CN104345115B (en) | A kind of method for qualitative and quantitative detection of Kanggu-Zengsheng tablet | |
| CN115032311B (en) | Method for measuring content of Xinnaoxin medicine components based on one-measurement-multiple-evaluation method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |