Method for detecting content of wild baicalin in chinese medicinal materials

Abstract

The present application is suitable for Scutellarein detection technical field, especially relates to a method for detecting the content of Scutellarein in traditional Chinese medicinal materials, comprising the following steps: step S1: selecting a predetermined amount of the to-be-tested traditional Chinese medicinal material, crushing it, mixing it with a predetermined amount of quartz sand, and then using a first pre-set concentration of ethanol solution for first extraction to obtain a preliminary extraction solution; step S2: performing solid-liquid separation on the preliminary extraction solution to separate out the ethanol extraction solution and retain the dregs; step S3: weighing a certain amount of the dregs, using a second pre-set concentration of ethanol solution for second extraction, and taking a proper amount of the second extraction solution as a sample solution. The present application realizes efficient and accurate detection of Scutellarein by optimizing the extraction conditions and the liquid chromatography detection steps. First, 70% ethanol solution is used for first extraction, which effectively dissolves Scutellarein in the traditional Chinese medicinal material, reduces impurity interference, and improves the purity of the preliminary extraction solution.

Description

Technical Field

[0001] This invention relates to the field of baicalin detection technology, specifically a method for detecting the content of baicalin in traditional Chinese medicinal materials. Background Technology

[0002] Baicalin is a natural compound with diverse pharmacological activities, widely found in the traditional Chinese medicine Scutellaria baicalensis and related plants. Due to its anti-inflammatory, antibacterial, and antiviral effects, it has become an important component for quality control in traditional Chinese medicine. Currently, the detection of baicalin mainly relies on high-performance liquid chromatography (HPLC), ultraviolet-visible spectrophotometry (UV-Vis), and enzyme-linked immunosorbent assay (ELISA). Among these, HPLC, due to its high sensitivity and high resolution, has become the primary method for detecting baicalin content. Traditional HPLC detection methods typically require complex sample pretreatment, including lengthy solvent extraction, liquid-liquid partitioning, and multiple centrifugation operations. While these steps effectively extract the target component, they also make the detection process cumbersome and increase sample loss and detection costs.

[0003] In existing technologies, extraction using a single ethanol concentration is common. However, this method is prone to incomplete extraction of some target components, especially when the residue contains high levels of baicalin, making it difficult to fully recover the active ingredients in a single extraction. Furthermore, multiple centrifugation operations are time-consuming and carry the risk of sample loss, hindering rapid processing of batches of samples. In addition, traditional methods often employ simple filtration, which fails to effectively remove fine suspended particles, easily causing column clogging and affecting the stability and accuracy of the detection results. Therefore, while existing technologies can meet basic detection requirements, they still have shortcomings in sample processing speed, extraction efficiency, and detection stability. Summary of the Invention

[0004] This invention provides the following technical solution:

[0005] The method for detecting the content of baicalin in traditional Chinese medicinal materials includes the following steps:

[0006] Step S1: Select a predetermined amount of Chinese medicinal materials to be tested, crush them, add a quantitative amount of quartz sand and mix, and then use an ethanol solution of the first preset concentration for the first extraction to obtain a preliminary extraction solution;

[0007] Step S2: Perform solid-liquid separation on the preliminary extraction solution to separate the ethanol extract and retain the residue;

[0008] Step S3: Weigh a certain amount of the residue, use a second preset concentration of ethanol solution for secondary extraction, and take an appropriate amount of the secondary extraction solution as the sample solution.

[0009] Step S4: The sample solution obtained from the secondary extraction is subjected to ultrasonic and filtration treatment, and the sample solution is cooled after filtration.

[0010] Step S5: Inject the cooled sample solution into the liquid chromatograph and determine the content of baicalin. The liquid chromatographic conditions are: acetonitrile-methanol-0.1% phosphoric acid with a mobile phase volume ratio of 11.5:12:70, column temperature of 35℃, and detection wavelength of 400nm.

[0011] Furthermore, the first preset concentration of ethanol solution used in step S1 is a 70% ethanol solution.

[0012] Furthermore, in step S2, solid-liquid separation is performed using a high-speed centrifugation method at a speed of 4000 rpm for 10 minutes.

[0013] Furthermore, the second preset concentration of ethanol solution used in step S3 is a 60% ethanol solution.

[0014] Furthermore, in step S4, when the sample solution is sonicated, the ultrasonic power is 200W and the duration is 5 minutes.

[0015] Furthermore, in step S4, the sample solution is filtered using a 0.45 μm filter membrane.

[0016] Furthermore, the detection conditions for liquid chromatography in step S5 include using acetonitrile-methanol-0.1% phosphoric acid in a volume ratio of 11.5:12:70 as the mobile phase, with a flow rate of 1.0 mL / min.

[0017] The method for detecting the content of baicalin in traditional Chinese medicinal materials provided by this invention has the following beneficial effects:

[0018] By optimizing extraction conditions and liquid chromatography detection steps, efficient and accurate detection of baicalin was achieved. First, a primary extraction using 70% ethanol solution effectively dissolves baicalin in the medicinal material while reducing interference from impurities and improving the purity of the initial extraction solution. Next, high-speed centrifugation achieves rapid and thorough solid-liquid separation, reducing sample loss and improving extraction efficiency. A secondary extraction using 60% ethanol solution further extracts residual baicalin from the residue, ensuring full recovery of the active ingredient and significantly improving detection sensitivity. Ultrasonic treatment accelerates component dissolution, and filtration through a 0.45 μm membrane ensures the clarity of the sample solution. The optimized liquid chromatography conditions ensure good separation and accurate detection of baicalin. Compared with existing technologies, this invention's method is simple to operate, rapid, and highly sensitive, making it particularly suitable for quality control and large-scale sample screening of medicinal materials, and has promising application prospects. Detailed Implementation

[0019] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.

[0020] The specific implementation of the present invention will be described in detail below with reference to specific embodiments.

[0021] The method for detecting the content of baicalin in traditional Chinese medicinal materials provided in this invention is characterized by comprising the following steps:

[0022] Step S1: Select a predetermined amount of Chinese medicinal materials to be tested, crush them, add a quantitative amount of quartz sand and mix, and then use an ethanol solution of the first preset concentration for the first extraction to obtain a preliminary extraction solution;

[0023] Step S2: Perform solid-liquid separation on the preliminary extraction solution to separate the ethanol extract and retain the residue;

[0024] Step S3: Weigh a certain amount of the residue, use a second preset concentration of ethanol solution for secondary extraction, and take an appropriate amount of the secondary extraction solution as the sample solution.

[0025] Step S4: The sample solution obtained from the secondary extraction is subjected to ultrasonic and filtration treatment, and the sample solution is cooled after filtration.

[0026] Step S5: Inject the cooled sample solution into the liquid chromatograph and determine the content of baicalin. The liquid chromatographic conditions are: acetonitrile-methanol-0.1% phosphoric acid with a mobile phase volume ratio of 11.5:12:70, column temperature of 35℃, and detection wavelength of 400nm.

[0027] In one embodiment of the present invention, firstly, a predetermined amount of the Chinese medicinal material to be tested is selected and pulverized into a fine powder to increase the surface area of ​​the sample, thereby improving the contact efficiency of the solvent. Then, the pulverized sample is weighed and a measured amount of quartz sand is added and thoroughly mixed. This prevents the sample from clumping during subsequent extraction and ensures the uniformity of the extraction process. Next, the mixed sample is subjected to a first extraction using an ethanol solution of a first preset concentration. Ultrasonic-assisted extraction can significantly improve the dissolution rate of baicalin, yielding a preliminary extraction solution.

[0028] In step S2, the preliminary extraction solution is subjected to solid-liquid separation. High-speed centrifugation can quickly and effectively separate the ethanol extract from the solid residue. This method can reduce sample loss during the separation process and improve the purity of the extract, which is beneficial for subsequent processing.

[0029] Next, in step S3, a certain amount of the residue is weighed and subjected to a second extraction using a second preset concentration of ethanol solution. The second extraction is also ultrasound-assisted, which can further extract residual baicalin from the residue, ensuring the full recovery of the active ingredients. An appropriate amount of the second extraction solution is used as the sample solution. This second extraction improves the comprehensiveness of the extraction, thereby making the measurement results more accurate.

[0030] In step S4, the obtained secondary extraction sample solution is ultrasonically treated and then filtered through a 0.45 μm filter membrane to effectively remove tiny particles from the sample and avoid interference in subsequent analysis. The filtered sample solution is then cooled to 4°C to reduce the loss of volatile components and maintain the stability of the sample components, ensuring the accuracy of the measurement results.

[0031] In step S5, the cooled sample solution is injected into the liquid chromatograph. The liquid chromatographic conditions are set as follows: mobile phase volume ratio of acetonitrile-methanol-0.1% phosphoric acid (11.5:12:70), column temperature controlled at 35℃, and detection wavelength at 400nm. This mobile phase ratio and chromatographic conditions ensure good separation and detection sensitivity of baicalin, thereby enabling accurate determination of the baicalin content in the sample.

[0032] This method, employing ultrasound-assisted extraction, high-speed centrifugation, secondary extraction, and optimized liquid chromatography conditions, achieves rapid and accurate detection of baicalin content in traditional Chinese medicinal materials. The secondary extraction step significantly improves the extraction efficiency of residual baicalin in the sample. By reusing ethanol solutions of different concentrations, incompletely extracted baicalin from the initial extraction can be effectively recovered, contributing to higher total extraction yields and more comprehensive analytical results. This secondary extraction step not only ensures the full recovery of active ingredients but also improves the accuracy of the determination results, reducing errors caused by residual active ingredients in the sample. Compared to existing technologies, this invention simplifies the operation process, reduces sample processing time, and improves detection sensitivity and accuracy, making it particularly suitable for quality control and large-scale sample screening of traditional Chinese medicinal materials, especially for the detection of trace components in complex matrix samples.

[0033] In this embodiment, the first preset concentration of ethanol solution used in step S1 is a 70% ethanol solution. Using a 70% ethanol solution helps improve the solubility of baicalin during the initial extraction, because 70% ethanol possesses both polar and non-polar solvent properties, enabling more effective dissolution and extraction of baicalin and other components from the medicinal material. This concentration balance ensures good extraction efficiency while reducing the dissolution of impurities caused by water, resulting in a purer extract and providing a better foundation for subsequent separation and detection.

[0034] In this embodiment, during solid-liquid separation in step S2, high-speed centrifugation is used at a speed of 4000 rpm for 10 minutes. High-speed centrifugation can quickly and efficiently separate the ethanol extract from the solid residue. The strong centrifugal force generated by high-speed centrifugation promotes the rapid sedimentation of small particles, significantly improving the clarity of the extract. It also reduces residual ethanol extract in the residue, avoiding cross-contamination during subsequent secondary extraction and improving sample recovery and purity.

[0035] In this embodiment, the second preset concentration of ethanol solution used in step S3 is a 60% ethanol solution. A 60% ethanol solution allows for better secondary extraction of baicalin from the residue. The lower polarity allows for more effective dissolution of residual baicalin in the residue, thereby further improving the extraction rate. Simultaneously, this concentration avoids the problem of excessive dissolution of other impurities caused by higher concentrations of ethanol, thus improving the purity of the sample solution and providing a more stable sample matrix for subsequent detection.

[0036] In this embodiment, during the ultrasonic treatment of the sample solution in step S4, the ultrasonic power is 200W and the duration is 5 minutes. This ultrasonic treatment can effectively promote particle dispersion and component dissolution in the sample solution within a short time. The ultrasonic waves can disrupt cell walls, accelerate the release of baicalin, and avoid sample degradation that may be caused by prolonged ultrasonic treatment. This ensures extraction efficiency while reducing the loss of effective components, thereby improving the efficiency of the extraction process and the quality of the sample.

[0037] In this embodiment, in step S4, a 0.45 μm filter membrane is used to filter the sample solution. The 0.45 μm filter membrane effectively removes fine particles and suspended solids from the sample solution, preventing these minute impurities from interfering with subsequent liquid chromatography analysis. The sufficiently small pore size of this filter membrane ensures the clarity of the sample solution without significantly affecting the recovery rate of the target compound, thus providing a more stable and pure sample solution for liquid chromatography analysis and guaranteeing the accuracy of the determination results.

[0038] In this embodiment, the detection conditions for liquid chromatography in step S5 include using acetonitrile-methanol-0.1% phosphoric acid in a volume ratio of 11.5:12:70 as the mobile phase, with a flow rate of 1.0 mL / min. This mobile phase ratio provides stable flow rate and pressure conditions while ensuring good separation, which helps optimize the separation between baicalin and other components, resulting in more symmetrical chromatographic peaks and higher detection sensitivity. The flow rate of 1.0 mL / min allows for sample detection within a reasonable time, ensuring experimental efficiency and data reproducibility.

[0039] The following are the experimental results based on the technical solutions mentioned above.

[0040] Experimental Report: Detection Method for the Content of Scutellarin in Chinese Medicinal Herbs

[0041] Experimental Background: Scutellarin is an important active ingredient in traditional Chinese medicine, widely used in the research of pharmacological effects such as anti-inflammatory and antibacterial, and in the quality control of traditional Chinese medicine. In order to achieve rapid and accurate determination of the content of scutellarin in Chinese medicinal herbs, a detection method based on ethanol extraction and liquid chromatography analysis was developed. By optimizing the extraction, separation and detection conditions, the detection efficiency and precision were improved.

[0042] Experimental Objectives:

[0043] 1. Optimize the pretreatment method of Chinese medicinal herb samples to improve the extraction efficiency of scutellarin.

[0044] 2. Determine the optimal liquid chromatography detection conditions to ensure high-sensitivity detection of scutellarin.

[0045] 3. Evaluate the effects of different extraction and separation conditions on sample purity and detection precision.

[0046] Materials and Methods:

[0047]

[0048] Experimental Results and Analysis:

[0049]

[0050] The R 2 value of the standard curve is 0.999, indicating that the detection method has a good linear relationship and is suitable for the quantitative analysis of scutellarin. The relative standard deviation (RSD) is all lower than 3%, indicating that the experimental method has good repeatability. According to the pharmacopoeia standard, the content of scutellarin in the samples is qualified when it is between 0.90 - 1.50 mg / g, and all samples meet the standard. Conclusion: In this experiment, by optimizing the primary extraction with 70% ethanol and the secondary extraction with 60% ethanol, the efficient extraction of scutellarin in Chinese medicinal herbs was achieved, significantly improving the extraction rate of scutellarin and the purity of the sample solution. By using high-speed centrifugation and ultrasonic treatment, the solid-liquid separation efficiency and extraction effect were improved, reducing the impurity content in the samples and providing a pure sample solution for subsequent liquid chromatography detection.

Claims

1. A method for detecting the content of baicalin in traditional Chinese medicinal materials, characterized in that, Includes the following steps: Step S1: Select a predetermined amount of Chinese medicinal materials to be tested, crush them, add a quantitative amount of quartz sand and mix, then use an ethanol solution of the first preset concentration for the first extraction to obtain a preliminary extraction solution; Step S2: Perform solid-liquid separation on the preliminary extraction solution to separate the ethanol extract and retain the residue; Step S3: Weigh a certain amount of the residue, use a second preset concentration of ethanol solution for secondary extraction, and take an appropriate amount of the secondary extraction solution as the sample solution. Step S4: The sample solution obtained from the secondary extraction is subjected to ultrasonic and filtration treatment, and the sample solution is cooled after filtration. Step S5: Inject the cooled sample solution into the liquid chromatograph and determine the content of baicalin. The liquid chromatographic conditions are: acetonitrile-methanol-0.1% phosphoric acid with a mobile phase volume ratio of 11.5:12:70, column temperature of 35℃, and detection wavelength of 400nm.

2. The method for detecting the content of baicalin in traditional Chinese medicinal materials according to claim 1, characterized in that, The first preset concentration of ethanol solution used in step S1 is a 70% ethanol solution.

3. The method for detecting the content of baicalin in traditional Chinese medicinal materials according to claim 1, characterized in that, In step S2, solid-liquid separation is performed using high-speed centrifugation at a speed of 4000 rpm for 10 minutes.

4. The method for detecting the content of baicalin in traditional Chinese medicinal materials according to claim 1, characterized in that, The second preset concentration of ethanol solution used in step S3 is a 60% ethanol solution.

5. The method for detecting the content of baicalin in traditional Chinese medicinal materials according to claim 1, characterized in that, In step S4, when the sample solution is sonicated, the ultrasonic power is 200W and the duration is 5 minutes.

6. The method for detecting the content of baicalin in traditional Chinese medicinal materials according to claim 1, characterized in that, In step S4, the sample solution is filtered using a 0.45 μm filter membrane.

7. The method for detecting the content of baicalin in traditional Chinese medicinal materials according to claim 1, characterized in that, The detection conditions for liquid chromatography in step S5 include using acetonitrile-methanol-0.1% phosphoric acid in a volume ratio of 11.5:12:70 as the mobile phase, and a flow rate of 1.0 mL / min.

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