Method for identifying under-forest mountain ginseng and garden ginseng based on proportion of ginseng monomer saponin

Abstract

The invention relates to an under-forest mountain ginseng and garden ginseng identification method based on a ginsenoside proportion, and belongs to the field of traditional Chinese medicinal material identification methods. The method specifically comprises the following steps: determining the contents of ginsenosides Rg1, Re and Rb1 of ginseng monomers by adopting a high performance liquid chromatography, calculating the Rb1 / (Rg1 + Re) proportion characteristic, and identifying the understory ginseng and the garden ginseng. The method has the advantages of being small in sample dosage, simple, high in method precision and repeatability and the like, the contents of the three kinds of ginseng monomer saponin specified by the 2020 edition Chinese Pharmacopoeia are measured, the proportion of the three kinds of ginseng monomer saponin can be calculated to serve as an identification index of the understory ginseng and the garden ginseng, and it is stipulated that the proportion Rb1 / (Rg1 + Re) of the ginseng monomer saponin in the understory ginseng is larger than 0.6, and the proportion Rb1 / (Rg1 + Re) of the ginseng monomer saponin in the garden ginseng is smaller than 0.6.

Description

Technical Field

[0001] The present invention relates to the field of traditional Chinese medicine identification methods, and in particular to an identification method for wild ginseng and garden ginseng based on the ratio of ginseng monomer saponins. Background Art

[0002] Panax ginseng Panaxginseng The dried roots and rhizomes of CA Mey. "Under-forest ginseng" refers to ginseng grown naturally in the wild in the mountains, commonly known as "seed sea ginseng," while "garden ginseng" refers to cultivated ginseng. Under-forest ginseng grows in a similar environment to wild ginseng, but due to less human intervention during cultivation, yields are lower, yet it possesses higher economic and medicinal value.

[0003] Modern pharmacological research indicates that ginseng possesses diverse pharmacological activities, with its numerous chemical components closely related to its diverse pharmacological effects. Its main chemical components include ginsenosides, ginseng polysaccharides, and ginsenoside vinol. Currently, there is extensive research on ginsenosides, and the 2020 edition of the Chinese Pharmacopoeia specifies content limits for the monomeric ginsenosides Rg1, Re, and Rb1 under the ginseng section.

[0004] Currently, there are many studies on garden ginseng and forest ginseng, but there is no significant and effective method to achieve the effective and rapid identification of garden ginseng and forest ginseng. This is also a key issue affecting the development of forest ginseng industry. Therefore, research on the identification of forest ginseng and garden ginseng is urgently needed. Summary of the Invention

[0005] In response to the problems existing in the prior art, the present invention provides a method for identifying wild ginseng grown in forests and garden ginseng based on the ratio of ginsenoside monomers. The present invention has the advantages of small sample dosage, simplicity, high method precision and repeatability, and can achieve rapid and accurate identification of wild ginseng grown in forests and garden ginseng medicinal materials.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions.

[0007] The invention discloses a method for identifying wild ginseng and garden ginseng under forests, which is characterized in that the method is to determine the content and content ratio of ginsenoside monomers in ginseng samples as analysis indicators.

[0008] Furthermore, the ginsenoside ratio is the ratio of ginsenoside monomer Rb1 to the sum of the contents of ginsenoside monomers Rg1 and Re.

[0009] Furthermore, the content of each monomeric ginsenoside in the ginseng sample is determined by HPLC.

[0010] Furthermore, the ginsenoside monomers include ginsenoside Rg1, ginsenoside Re, and ginsenoside Rb1.

[0011] The present invention also discloses the application of any of the above identification methods in preparing products for forest ginseng and garden ginseng.

[0012] The present invention also discloses a method for identifying the above-mentioned wild ginseng and garden ginseng, the specific steps of which include: Step 1. Preparation of standard solution: Accurately weigh ginsenoside Rg1, ginsenoside Re, and ginsenoside Rb1 reference substances, add methanol to make a mixed solution containing 0.2 mg of each substance per 1 mL, and shake well. Step 2. Preparation of test solution: Take dried ginseng, crush it (pass it through a No. 4 sieve), weigh about 0.5 g of ginseng powder, accurately weigh it, add 10 mL of 70% methanol, weigh it, and ultrasonically extract it for 30 minutes. After cooling to room temperature, add 70% methanol to make up the loss, filter it, and take the filtrate to obtain the test solution; Step 3, determination of ginsenoside monomers: taking the sample to be tested in step 1, placing it in a high performance liquid chromatograph, and determining the content ratio of ginsenoside monomers; Step 4. HPLC is used to determine the peak area of each ginsenoside monomer in the sample and compare it with the monosaccharide peak area of the reference substance. The ginsenoside monomer content of the sample is calculated by the external standard method, and the ratio of ginsenoside monomer Rb1 to the sum of the contents of ginsenoside monomers Rg1 and Re is calculated (Rb1 / (Rg1+Re)).

[0013] Furthermore, the ratio of ginseng monomer saponin content is: the ratio of ginseng monomer saponin in wild ginseng is Rb1 / (Rg1+Re)>0.6, and the ratio of ginseng monomer saponin in garden ginseng is Rb1 / (Rg1+Re)<0.6.

[0014] The present invention also discloses the application of any of the above identification methods in preparing products for forest ginseng and garden ginseng.

[0015] The present invention also discloses a model for identifying undergrowth wild ginseng and garden ginseng, which is characterized in that it is constructed by any of the above-mentioned methods for identifying undergrowth wild ginseng and garden ginseng.

[0016] The present invention also discloses a device for identifying wild ginseng and garden ginseng under forest, which is characterized by comprising the wild ginseng and garden ginseng identification model described above.

[0017] Compared with the prior art, the present invention has the following beneficial effects.

[0018] The method for distinguishing wild ginseng from garden ginseng based on the ratio of ginsenoside monomers described in the present invention has not been reported in the prior art. Furthermore, the use of high-performance liquid chromatography (HPLC) using the ratio of ginsenoside monomers Rg1, Re, and Rb1 as a limit for distinguishing wild ginseng from garden ginseng is unprecedented. The present invention provides methods for extracting ginsenosides from wild ginseng and garden ginseng and for determining their content. This method is simple and easy to operate, has a simple and environmentally friendly process, and can effectively distinguish wild ginseng from garden ginseng, providing a theoretical basis for the development of new drugs for wild ginseng medicinal materials and powders. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 The HPLC spectra of the reference substances ginsenosides Rg1, Re, and Rb1 in the present invention are shown in Figure 1. Note: 1-Rg1; 2-Re; 3-Rb1.

[0020] Figure 2 This is the HPLC spectrum of the ginsenoside sample of the present invention. Note: 1-Rg1; 2-Re; 3-Rb1. DETAILED DESCRIPTION

[0021] The present invention is further described in detail below with reference to specific embodiments. However, this should not be construed as limiting the scope of the present invention to the following embodiments, as all technologies implemented based on the present invention fall within the scope of the present invention.

[0022] Unless otherwise specified, all reagents and materials used in the present invention are commercially available.

[0023] Example 1: Preparation of standard solution: Accurately weigh ginsenoside Rg1 reference substance, ginsenoside Re reference substance and ginsenoside Rb1 reference substance, add methanol to make a mixed solution containing 0.2 mg of each per 1 mL, and shake well to obtain the standard solution.

[0024] Preparation of the test solution: Take 0.5 g of ginseng powder (passed through No. 4 sieve), accurately weigh it, place it in a stoppered conical flask, add 10 mL of 70% methanol, weigh it, and ultrasonically extract it for 30 minutes. After cooling to room temperature, make up the loss with 70% methanol, filter it, and take the filtrate to obtain the test solution.

[0025] Chromatographic conditions: A Waters Symmetry Shield RP18 (4.6 × 250 mm, 5 μm) column was used with a diode array detector (DAD). The mobile phase consisted of 0.1% phosphoric acid (A) and acetonitrile (B) in a gradient elution (see Table 1). The flow rate was 1.0 mL / min, the column temperature was 25°C, the detection wavelength was 203 nm, and the injection volume was 10 μL. The theoretical plate number, calculated based on the ginsenoside Rg1 peak, should be no less than 6000.

[0026] Table 1 Mobile phase gradient elution program .

[0027] After testing 10 batches of wild ginseng from forests of different growth years and 10 batches of garden ginseng, the ratio of ginsenoside monomers Rb1 / (Rg1+Re) in wild ginseng from forests was 0.71±0.08, and the ratio of ginsenoside monomers Rb1 / (Rg1+Re) in garden ginseng was 0.49±0.05.

[0028] Table 2 Experimental results .

[0029] Example 2: Preparation of reference solution: Accurately weigh ginsenoside Rg1 reference substance, ginsenoside Re reference substance and ginsenoside Rb1 reference substance, add methanol to make a mixed solution containing 0.2 mg of each per 1 mL, and shake well to obtain the standard solution.

[0030] Preparation of test solution: Take about 1g of ginseng powder (passed through No. 4 sieve), accurately weigh it, wrap it with neutral filter paper, place it in a Soxhlet extractor, add chloroform and heat under reflux for 3h, discard the chloroform solution, evaporate the solvent from the residue, transfer it together with the filter paper into a 100mL conical flask, accurately add 50mL of water-saturated n-butanol, plug it, leave it overnight, ultrasonically treat it for 30min, filter it, discard the initial filtrate, accurately measure 25mL of the subsequent filtrate, place it in an evaporating dish, evaporate the residue to dryness, add methanol to dissolve it and transfer it to a 5mL volumetric flask, add methanol to the scale, shake it up, filter it, take the subsequent filtrate to obtain the test solution.

[0031] Chromatographic conditions: Octadecyl bonded silica gel as the packing material, water (A)-acetonitrile (B) as the mobile phase for gradient elution (see Table 3); flow rate 1.0 mL / min; column temperature 25°C; detection wavelength 203 nm; injection volume 10 μL. The theoretical plate number calculated based on the ginsenoside Rg1 peak should be no less than 6000.

[0032] Table 3 Mobile phase gradient elution program .

[0033] After testing 10 batches of wild ginseng from forests of different growth years and 10 batches of garden ginseng, the ratio of ginsenoside monomers Rb1 / (Rg1+Re) in wild ginseng from forests was 0.71±0.07, and the ratio of ginsenoside monomers Rb1 / (Rg1+Re) in garden ginseng was 0.48±0.05.

[0034] Table 4 Experimental results .

[0035] Example 3: Verification experiment.

[0036] Ten batches of samples were randomly selected and blind tested according to the conditions described in Example 1. The experimental data and results are as follows: According to the test results, among the 10 batches of samples, there were 4 wild ginsengs and 6 garden ginsengs, and the ratios of ginsenoside content in the samples were 0.72±0.09 and 0.50±0.06 respectively. This conclusion was consistent with the samples taken.

[0037] Table 5 Verification experimental results .

[0038] In summary, the methods provided herein for extracting ginsenosides and determining the content and ratio of ginsenoside monomers are highly effective in distinguishing wild ginseng from garden ginseng. Based on the assay results, the ratio of ginsenoside monomers, Rb1 / (Rg1+Re), is defined as greater than 0.6 in wild ginseng and less than 0.6 in garden ginseng. This method is simple, rapid, and environmentally friendly, thus laying the foundation for the further development and utilization of wild ginseng and possessing broad prospects.

[0039] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the scope of the present invention. Those skilled in the art will readily appreciate that the present invention is susceptible to various modifications and variations. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention shall be included within the scope of protection of the present invention.

Claims

1. A method for distinguishing wild ginseng from garden ginseng, characterized in that: The method is to determine the content and content ratio of ginsenoside monomers in ginseng samples as analytical indicators.

2. The identification method according to claim 1, characterized in that: The ginsenoside ratio is the ratio of ginsenoside monomer Rb1 to the sum of the contents of ginsenoside monomers Rg1 and Re.

3. The identification method according to claim 1, characterized in that: The content of each monomeric ginsenoside in the ginseng sample was determined by HPLC.

4. The identification method according to claim 3, characterized in that: The ginsenoside monomers include ginsenoside Rg1, ginsenoside Re, and ginsenoside Rb1.

5. Use of the identification method according to any one of claims 1 to 4 in the preparation of products for forest ginseng and garden ginseng.

6. A method for identifying wild ginseng and garden ginseng according to claim 1, comprising the following steps: Step 1. Preparation of standard solution: Accurately weigh ginsenoside Rg1, ginsenoside Re, and ginsenoside Rb1 reference substances, add methanol to make a mixed solution containing 0.2 mg of each substance per 1 mL, and shake well. Step 2. Preparation of test solution: Take dried ginseng, grind it (pass it through a No. 4 sieve), weigh about 0.5 g of ginseng powder, accurately weigh it, add 10 mL of 70% methanol, weigh it, and ultrasonically extract it for 30 minutes. After cooling to room temperature, add 70% methanol to make up the loss, filter it, and take the filtrate to obtain the test solution; Step 3, determination of ginsenoside monomers: taking the sample to be tested in step 1, placing it in a high performance liquid chromatograph, and determining the content ratio of ginsenoside monomers; Step 4. HPLC is used to determine the peak area of each ginsenoside monomer in the sample and compare it with the monosaccharide peak area of the reference substance. The ginsenoside monomer content of the sample is calculated by the external standard method, and the ratio of ginsenoside monomer Rb1 to the sum of the contents of ginsenoside monomers Rg1 and Re is calculated (Rb1 / (Rg1+Re)).

7. The identification method according to claim 5, characterized in that: The ratio of ginseng monomer saponin content is: the ratio of ginseng monomer saponin in wild ginseng is Rb1 / (Rg1+Re)>0.6, and the ratio of ginseng monomer saponin in garden ginseng is Rb1 / (Rg1+Re)<0.

6.

8. Use of the identification method according to any one of claims 6 to 7 in the preparation of products for forest ginseng and garden ginseng.

9. A model for identifying under-forest mountain ginseng and garden ginseng, characterized in that: The method for identifying undergrowth wild ginseng and garden ginseng is constructed according to any one of claims 5 to 6.

10. A device for identifying wild ginseng and garden ginseng, characterized in that: It includes the forest ginseng and garden ginseng identification model as described in claim 8.

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