CN111830143A - Method for measuring content of paeoniflorin in traditional Chinese medicine preparation - Google Patents
Method for measuring content of paeoniflorin in traditional Chinese medicine preparation Download PDFInfo
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- CN111830143A CN111830143A CN201910326978.XA CN201910326978A CN111830143A CN 111830143 A CN111830143 A CN 111830143A CN 201910326978 A CN201910326978 A CN 201910326978A CN 111830143 A CN111830143 A CN 111830143A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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Abstract
The invention discloses a method for measuring the content of paeoniflorin in a traditional Chinese medicine preparation, which comprises the following steps: using SHIMADZU shim-pack GIST C18 chromatographic column (5 μm 4.6 × 250mm), using 70% acetonitrile-0.1% -0.5% phosphoric acid water solution (when 0-20min, mobile phase A: mobile phase B is 20: 80-30: 7020-21min, mobile phase A: mobile phase B is 30: 70-100: 0) as fluidity, flow rate is 1.0m L min‑1And the detection wavelength is 230 nm. Through methodology investigation, the method has good system adaptability, specificity, linear relation and durability, is simple, convenient and quick, has accurate and reliable result, and can be used for measuring the content of paeoniflorin in the prostate capsule.
Description
Technical Field
The invention belongs to the technical field of traditional Chinese medicines, and particularly relates to a method for measuring the content of paeoniflorin in a traditional Chinese medicine preparation.
Background
The Qianlieantong capsule is prepared from 8 traditional Chinese medicines of golden cypress, red paeony root, red-rooted salvia root, peach seed, hiraute shiny bugleweed herb, combined spicebush root, cowherb seed and dahurian angelica root, and has the functions of clearing heat and promoting diuresis, and activating blood and dissolving stasis. Can be used for treating syndrome of damp-heat stagnation, manifested by frequent micturition, urgent micturition, difficulty in micturition, and lower abdominal distention and pain. Wherein radix Paeoniae Rubra and cortex Phellodendri are used as principal drugs, and their effective components are penoniflorin and berberine hydrochloride respectively. The medicine mainly contains paeoniflorin, berberine hydrochloride and salvianolic acid B. As for the paeoniflorin content determination method, a thin-layer scanning method, a capillary electrophoresis method, a thin-layer-ultraviolet spectrophotometry method, a high performance liquid chromatography method and the like are reported, but in the actual production detection process, the existing detection method has poor repeatability of the paeoniflorin content and has errors. In view of the above reasons, the invention provides a detection method of the Qianlieantong capsule paeoniflorin, which has the advantages of simple operation, rapidness, accuracy, high sensitivity, good repeatability and good separation effect, and is convenient for the quality control of products.
Disclosure of Invention
The invention aims to overcome the defects of the prior art, and provides a detection method of Qianlieantong capsule paeoniflorin, namely a detection method of Qianlieantong capsule paeoniflorin.
The purpose of the invention is realized by the following technical scheme:
a method for measuring paeoniflorin in Qianlieantong capsules comprises the following steps:
(1) preparation of assay solution: taking contents of the Qianlieantong capsule, grinding, taking 0.3g, precisely weighing, placing in a conical flask with a plug, precisely adding 25ml of methanol, weighing, soaking for 4 hours, ultrasonically treating for 20 minutes, cooling, weighing again, supplementing the lost weight with methanol, shaking up, filtering, and taking the subsequent filtrate as a test solution; taking appropriate amount of penoniflorin as control, precisely weighing, and adding methanol to obtain solution containing 0.1mg per 1ml as control solution.
(2) Determination of the solution: sucking the test solution and the reference solution, injecting into a liquid chromatograph, and performing high performance liquid chromatography by gradient elution to obtain chromatogram of the test solution and penoniflorin reference solution of the Qianlieantong capsule;
wherein, the chromatographic conditions of the high performance liquid chromatography are as follows: the filler of the chromatographic column is octadecylsilane chemically bonded silica; the mobile phase A is an aqueous solution containing 60-100% of acetonitrile, the mobile phase B is an aqueous solution containing 0.1-0.5% of phosphoric acid or acetic acid, and the volume ratio of the mobile phase A to the mobile phase B in the mobile phase is gradually increased along with the increase of time.
The detection wavelength is 200-300nm, and the detection wavelength of the high performance liquid chromatography is preferably 230 nm.
The temperature of the chromatographic column is 30-50 ℃, and preferably 40 ℃; the mobile phase A is an aqueous solution containing 60-100% of acetonitrile, and preferably the acetonitrile in the mobile phase A accounts for 70%; the mobile phase B contains 0.1-0.5% phosphoric acid or acetic acid aqueous solution, preferably 0.1% phosphoric acid aqueous solution.
The procedure for gradient elution by HPLC is as follows, wherein the following proportions are by volume:
when the time is 0-20min, the ratio of mobile phase A to mobile phase B is 20: 80-30: 70
At 20-21min, the ratio of mobile phase A to mobile phase B is 30: 70-100: 0
The following gradient elution procedure is preferred:
at 0min, the ratio of mobile phase A to mobile phase B is 20: 8
At 20min, the ratio of mobile phase A to mobile phase B is 30: 70
At 25min, the ratio of mobile phase A to mobile phase B is 100: 0
The invention provides a method for measuring the content of paeoniflorin in a Qianlieantong capsule. The method is a feasible method obtained on the basis of a large number of experiments, and has repeatability. In the invention, the paeoniflorin content in the same batch of Qianlieantong capsules obtained under the same test condition is basically the same through high performance liquid chromatography, which indicates that the repeatability is good. The paeoniflorin recovery rate is measured by adding a paeoniflorin reference substance, the recovery rate is more than 95 percent, and the method is proved to have good accuracy. Therefore, the determination method established by the determination method can be used as the quality control standard of the prostate capsule, and the method can objectively reflect the content of paeoniflorin in the product and can more comprehensively control the quality of the product.
The invention has the following advantages:
(1) the method has the characteristics of simplicity, stability, high precision, good reproducibility and easy control.
(2) Can be used for the investigation of the stability of the production process and the stability of the product, and is beneficial to monitoring the quality of the preparation more comprehensively.
Drawings
FIG. 1 is a chromatogram of a paeoniflorin control.
FIG. 2 is a chromatogram of the Qianlieantong capsule sample.
FIG. 3 is HPLC chromatogram of Qianlieantong capsule lacking radix Paeoniae Rubra sample
Detailed Description
The following detailed description of the embodiments of the present invention is provided for the purpose of illustration and not limitation, and should not be construed as limiting the scope of the invention.
Example 1
1. Instruments and reagents
The instrument comprises the following steps: an Agilent 1260 Infinity II high performance liquid chromatograph equipped with an ultraviolet detector; acetonitrile and phosphoric acid are both chromatographically pure, and methanol and ethanol are analytically pure; ouhaha purified water; 11 batches of Qianlieantong capsule finished product.
2. Chromatographic conditions
SHIMADZU shim-pack GIST C185 μm 4.6 × 250 mm; the column temperature is 40 ℃; the flow rate is 1 ml/min; mobile phase a was 70% acetonitrile; mobile phase B was 0.1% phosphoric acid solution, wavelength: 230nm, gradient elution, elution program:
at 0min, the ratio of mobile phase A to mobile phase B is 20: 80;
at 20min, the ratio of mobile phase A to mobile phase B is 30: 70;
at 21min, mobile phase A and mobile phase B are 100: 0.
3. Preparation of samples
3.1 preparation of test solution:
3.1.1 preparation of the prostate capsule test solution; grinding the contents of the Qianlieantong capsule, precisely weighing 0.3g, placing in a conical flask with a plug, adding diluted ethanol 25ml, weighing, ultrasonic treating for 30min, cooling, supplementing weight, and filtering.
3.1.2 preparation of penoniflorin reference solution: taking 10mg of paeoniflorin reference substance, placing in a 10ml volumetric flask, adding methanol to dissolve and diluting to scale to obtain the final product.
3.1.3 preparation of prostate capsule negative sample solution: taking a red peony root sample of the Qianlieantong capsule, grinding, taking 0.3g, precisely weighing, placing in a conical flask with a plug, adding 25ml of diluted ethanol, weighing, carrying out ultrasonic treatment for 30min, cooling, supplementing weight, and filtering to obtain the medicine.
3.2 assay: respectively sucking 10ul of the test solution and the reference solution, respectively, injecting into a liquid chromatograph for measurement, and recording the chromatogram. And calculating the content of paeoniflorin by a peak area according to an external standard method.
The method specifically comprises the following steps:
3.2.1 FIG. 1 is the chromatogram of paeoniflorin control.
3.2.2 FIG. 2 is the chromatogram of the Qianlieantong capsule sample.
3.3 methodological inspection
3.3.1 System applicability
And continuously feeding the same paeoniflorin reference substance solution for 6 times, observing the system applicability of the instrument, and calculating by using the retention time and the peak area of the main peak of the paeoniflorin, wherein the retention time RSD (%) is 0.40%, the peak area RSD (%) is 0.42%, and the minimum value of the theoretical plate number is 17070. The results show that the system is good in applicability.
3.3.2 specialization examination
Collecting paeoniflorin reference substance, negative sample solution, and sample solution, injecting into liquid chromatograph, inspecting method specificity, keeping the retention time of the main peak of the reference substance and the main peak of the sample solution consistent, and the negative sample does not interfere with the main peak detection. Indicating that the method has good specificity.
3.3.3 Linear relationship investigation
Taking a proper amount of reference substances, respectively preparing reference substance solutions with different concentrations, injecting the reference substance solutions into a liquid chromatograph, and performing linear regression on the concentrations by using the measured peak areas. The regression equation is that y is 12730x-5.487, and the correlation coefficient R20.9999. Indicating a good linear relationship.
3.3.4 repeatability test
Taking 6 parts of the same batch of Qianlieantong capsules, preparing the capsule according to the method under the item of the sample preparation method '3.1.1', and measuring, wherein the RSD (%) value of the paeoniflorin content is 0.97 percent by the external standard method and the peak area. The results show that the method has good repeatability.
3.3.5 accuracy test
Taking the Qianlieantong capsule, precisely adding a proper amount of paeoniflorin reference substance, preparing a test solution and a reference substance solution according to a method, measuring the content, wherein the average recovery rate of 9 samples of high, medium and low is 96.07, and the RSD (%) value of the recovery rate is 0.87%. The method has good accuracy.
3.3.6 durability examination
The durability of the chromatographic conditions was examined by slightly varying the flow rate, the ratio of mobile phases, the column temperature, the wavelength, the column, and other factors. The RSD (%) value of paeoniflorin content was 1.31% after slight variation of the chromatographic conditions. The method has good durability.
4. Ten batches of Qianlieantong capsule samples were determined
4.1 the content of paeoniflorin in ten batches of finished prostate capsule products is respectively determined according to the detection method and conditions.
5. Measurement results
5.1 the results of the paeoniflorin content in ten batches of the prostate capsule are respectively as follows: 2.66 mg/pellet, 2.60 mg/pellet, 2.48 mg/pellet, 3.05 mg/pellet, 2.35 mg/pellet, 3.51 mg/pellet, 3.48 mg/pellet, 2.85 mg/pellet, 4.08 mg/pellet, 2.98 mg/pellet.
Claims (1)
1. A method for measuring the content of paeoniflorin, a traditional Chinese medicine preparation, is characterized by comprising the following steps:
(1) preparation of assay solution: taking contents of the Qianlieantong capsule, grinding, taking 0.3g, precisely weighing, placing in a conical flask with a plug, precisely adding 25ml of methanol, weighing, soaking for 4 hours, ultrasonically treating for 20 minutes, cooling, weighing again, supplementing the lost weight with methanol, shaking up, filtering, and taking the subsequent filtrate as a test solution; taking a proper amount of paeoniflorin reference substance, precisely weighing, and adding methanol to obtain a solution containing 0.1mg per 1ml as reference substance solution;
(2) determination of the solution: sucking the test solution and the reference solution, injecting into a liquid chromatograph, and performing high performance liquid chromatography by gradient elution to obtain chromatogram of the test solution and penoniflorin reference solution of the Qianlieantong capsule;
wherein, the chromatographic conditions of the high performance liquid chromatography are as follows: the filler of the chromatographic column is octadecylsilane chemically bonded silica; the mobile phase A is an aqueous solution containing 60-100% of acetonitrile, and preferably the acetonitrile in the mobile phase A accounts for 70%; the mobile phase B is a phosphoric acid or acetic acid aqueous solution containing 0.1 to 0.5 percent, preferably a 0.1 percent phosphoric acid aqueous solution; the volume ratio of the mobile phase A to the mobile phase B in the mobile phase gradually increases along with the increase of time;
the detection wavelength is 200-300nm, preferably 230 nm;
the procedure for gradient elution by HPLC is as follows, wherein the following proportions are by volume:
when the time is 0-20min, the ratio of mobile phase A to mobile phase B is 20: 80-30: 70,
when 20-21min, the ratio of mobile phase A to mobile phase B is 30: 70-100: 0;
the following gradient elution procedure is preferred:
at 0min, the ratio of mobile phase A to mobile phase B is 20: 8
At 20min, the ratio of mobile phase A to mobile phase B is 30: 70
At 25min, the ratio of mobile phase A to mobile phase B is 100: 0
Precisely measuring 10 μ l of each of the reference solution and the sample solution, injecting into a liquid chromatograph according to chromatographic conditions, recording peak area, calculating content according to an external standard method, and measuring.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115420816A (en) * | 2022-07-21 | 2022-12-02 | 广东万年青制药股份有限公司 | Content determination method of traditional Chinese medicine composition for nourishing yin and cooling blood |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1818637A (en) * | 2006-03-20 | 2006-08-16 | 成都优他制药有限责任公司 | Quality inspection of Qianlieantong oral preparation |
| CN108195967A (en) * | 2018-01-10 | 2018-06-22 | 广西壮族自治区食品药品检验所 | The content assaying method of Paeoniflorin and calycosin glucoside in Chinese medicine preparation |
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- 2019-04-16 CN CN201910326978.XA patent/CN111830143A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1818637A (en) * | 2006-03-20 | 2006-08-16 | 成都优他制药有限责任公司 | Quality inspection of Qianlieantong oral preparation |
| CN108195967A (en) * | 2018-01-10 | 2018-06-22 | 广西壮族自治区食品药品检验所 | The content assaying method of Paeoniflorin and calycosin glucoside in Chinese medicine preparation |
Non-Patent Citations (3)
| Title |
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| ZHOU, XIAOYING 等: "Rapid determination of paeoniflorin from Paeonia sinjiang K. Y. Pan. by rapid resolution liquid chromatography", 《PHARMACOGNOSY MAGAZINE》 * |
| 丁仕强: "HPLC测定前列安通胶囊中芍药苷的含量", 《中国实验方剂学杂志》 * |
| 伦立军 等: "HPLC法测定前列欣胶囊中芍药苷的含量", 《齐鲁药事》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115420816A (en) * | 2022-07-21 | 2022-12-02 | 广东万年青制药股份有限公司 | Content determination method of traditional Chinese medicine composition for nourishing yin and cooling blood |
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Application publication date: 20201027 |