Method for determining artemisinin content in artemisia apiacea through rapid thin-layer analysis, application and colorimetric card

Abstract

The invention provides a method for determining the content of artemisinin in artemisia apiacea through rapid thin-layer analysis, application and a colorimetric card, and belongs to the technical field of substance detection. The method provided by the invention comprises the following steps: S1, mixing a to-be-detected medicinal material with a solvent, performing ultrasonic extraction, and filtering to obtain a to-be-detected solution; s2, the to-be-detected liquid obtained in the step S1 is subjected to sample application, unfolding and drying on a thin-layer plate, and a chromatography plate is obtained; s3, carrying out color development on the chromatography plate obtained in the step S2 to obtain a color-developed chromatography plate; s4, comparing and identifying the chromogenic chromatography plate obtained in the step S3 with a standard colorimetric card, and judging the value range of the artemisinin content. The method can be used for rapidly determining whether the artemisinin content in the sweet wormwood herb is qualified or not.

Description

Technical Field

[0001] The invention belongs to the technical field of material detection, and relates to a method for rapid thin-layer analysis to determine the content of artemisinin in artemisia annua, an application thereof and a colorimetric card. Background Art

[0002] Artemisinin is a small molecule drug derived from plants. Artemisinin has good efficacy for cerebral malaria and chloroquine-resistant malaria, and is fast-acting and low-toxic. Its anti-malarial mechanism of action is mainly that in the process of treating malaria, artemisinin activates free radicals, which bind to malarial proteins and act on the membrane structure of malarial parasites, causing damage to their vesicle membranes, nuclear membranes, and plasma membranes, swelling of mitochondria, and shedding of inner and outer membranes, thereby damaging the cell structure and function of malarial parasites, and affecting the chromatin in the nucleus to a certain extent. Artemisinin can also significantly reduce the intake of isoleucine by malarial parasites, thereby inhibiting the synthesis of parasite proteins. In recent years, the anti-tumor effect of artemisinin has gradually been discovered. Artemisinin can cause apoptosis of breast cancer cells, liver cancer cells, cervical cancer cells and other cancer cells, and has a significant inhibitory effect on the growth of cancer cells. The anti-tumor effect of artemisinin and its derivatives is mainly achieved by inducing cell apoptosis. Artemisia annua is an important natural source of artemisinin, and the rapid detection of artemisinin in artemisia annua is of great significance.

[0003] Chinese invention patent CN117191976A provides a method for detecting the content of deoxyartemisinin in Artemisia annua, comprising the following steps: S1, preparing a reference solution: taking deoxyartemisinin as a reference, preparing a reference solution; S2, preparing a test solution: taking the Artemisia annua sample to be tested as a test sample, preparing a test solution; S3, injecting the reference solution and the test solution into a gas chromatograph respectively to obtain a chromatogram; then according to the chromatograms of the reference and test samples, the content of deoxyartemisinin in the test solution is calculated. This technology provides a method for detecting the content of deoxyartemisinin in Artemisia annua by gas chromatography, which has strong specificity, good operability, good accuracy, linearity, precision and durability, and can achieve rapid quantitative determination of the content of deoxyartemisinin in Artemisia annua.

[0004] Chinese invention patent CN115728408A provides a method for simultaneously determining seven artemisinin-related compounds in Artemisia annua, which comprises using ultra-high performance liquid chromatography-tandem mass spectrometry to detect the contents of artemisinin, artemisinin B, artemisinic acid, artemisinene, dihydroartemisinic acid, artemisinic aldehyde, and Amorpha-4,11-diene in Artemisia annua extract. The method is simple to operate, highly sensitive, and accurate. Both of the above methods are based on chromatographic instrument detection and require the assistance of chromatographic instruments to complete.

[0005] Chinese invention patent CN113176218A provides a method for rapid detection of artemisinin content. The method includes: determining the optimal absorption wavelength; determining and eliminating background interference; establishing a standard curve; processing and measuring samples. This detection method directly adopts a 95% ethanol water bath heating extraction method, which reduces the sample pretreatment steps, reduces the pretreatment time and random errors; in addition, by utilizing the time difference between the reaction of NaOH solution and non-artemisinin interfering substances, and NaOH solution and artemisinin at room temperature, the background interference is eliminated to the maximum extent. The results show that the detection results are consistent with the standard HPLC detection values, and no linear fitting correction is required.

[0006] Chinese invention patent CN117147744A provides a method for detecting the artemisinin content in Artemisia annua seeds, the method comprising the following steps: (1) crushing the raw material: crushing the dry leaves of Artemisia annua to a particle size of less than 0.5 mm; (2) ultrasonic extraction: adding to petroleum ether and ultrasonicating in a water bath; (3) concentration: using an instrument to concentrate; (4) re-dissolving: re-dissolving in an organic solvent; (5) sampling detection: using a chromatograph to detect the artemisinin content; the results are stable and reproducible, which solves the problems of long detection time, large sample demand, high operation risk and poor reproducibility of conventional methods.

[0007] However, the existing technology cannot provide a method that can quickly and accurately detect and identify whether the content of artemisinin in the medicinal material Artemisia annua is within a qualified range. Summary of the invention

[0008] In view of this, the prior art cannot provide a method for quickly and accurately detecting and identifying whether the content of artemisinin in the medicinal material Artemisia annua is within the acceptable range. The purpose of the present invention is to provide a method, application and colorimetric card for rapid thin layer analysis to determine the content of artemisinin in Artemisia annua.

[0009] To achieve the above-mentioned object of the invention, on the one hand, the present invention provides a method for determining the content of artemisinin in Artemisia annua by rapid thin layer analysis, comprising the following steps: S1. Mix the medicinal material to be tested with the solvent, perform ultrasonic extraction, and filter to obtain the test solution; S2, spotting the test solution obtained in step S1 on the thin layer plate, developing, and drying to obtain a chromatography plate; S3, coloring the chromatography plate obtained in step S2 to obtain a color-developing chromatography plate; S4, comparing the color chromatography plate obtained in step S3 with a standard colorimetric card to determine the value range of the artemisinin content; Wherein, the standard colorimetric card in step S4 includes color block 1, color block 2, color block 3 and color block 4; The CMYK values ​​of the color block 1 are C=0, M=85, Y=70, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 1 is 16-20 mg / g; The CMYK values ​​of the color block 2 are C=0, M=65, Y=50, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 2 is 12-15 mg / g; The CMYK values ​​of the color block 3 are C=0, M=45, Y=30, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 3 is 7-11 mg / g; The CMYK values ​​of the color block 4 are C=0, M=20, Y=10, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 4 is 3-6 mg / g.

[0010] Preferably, in step S1, the solvent is at least one of petroleum ether, ethyl acetate and dichloromethane; and the ultrasonic extraction time is 30-60 min.

[0011] Preferably, in step S2, the thin layer plate is a silica gel G thin layer plate, and the developing agent is a mixture of petroleum ether and ethyl acetate in a volume ratio of 5-15:1.

[0012] Preferably, in step S2, the development is specifically as follows: placing the spotted silica gel G thin layer plate into a development cylinder, and the spotting origin cannot be immersed in the developing agent; when the front edge of the developing agent rises to the upper edge of the silica gel G thin layer plate, 10%-30% away from the spotting origin, the silica gel G thin layer plate is taken out.

[0013] Preferably, in step S3, the color development comprises the following steps: spraying the chromatography plate obtained in step S2 with a vanillin sulfate ethanol solution, and heating at 100-110° C. for 10-20 min.

[0014] Preferably, the vanillin sulfuric acid ethanol solution has a mass composition of: 1%-5% concentrated sulfuric acid, 5%-20% vanillin, and the remainder being ethanol.

[0015] More preferably, the mass composition of the vanillin sulfuric acid ethanol solution is: 2.5% concentrated sulfuric acid, 15% vanillin, and the balance is ethanol.

[0016] Preferably, in step S4, the comparative identification is specifically performed by placing the sample at room temperature and then comparing the sample with a standard colorimetric card under white light.

[0017] On the other hand, the present invention provides the use of the above method in determining the content of artemisinin.

[0018] In another aspect, the present invention provides a standard colorimetric card for use in the process of determining the content of artemisinin in Artemisia annua in the above method; The standard colorimetric card includes color block 1, color block 2, color block 3 and color block 4; The CMYK values ​​of the color block 1 are C=0, M=85, Y=70, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 1 is 16-20 mg / g; The CMYK values ​​of the color block 2 are C=0, M=65, Y=50, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 2 is 12-16 mg / g; The CMYK values ​​of the color block 3 are C=0, M=45, Y=30, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 3 is 7-12 mg / g; The CMYK values ​​of the color block 4 are C=0, M=20, Y=10, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 4 is 3-7 mg / g.

[0019] Compared with the prior art, the present invention has the following beneficial effects: The present invention provides a convenient, simple and easy-to-operate method for determining the content of artemisinin in Artemisia annua by rapid thin-layer analysis. The method is convenient for rapid semi-quantitative detection of artemisinin in large quantities of Artemisia annua medicinal materials, improves detection efficiency and saves detection costs. DETAILED DESCRIPTION

[0020] Terms and Claims of the Present Invention: 1. The articles “a”, “an” and “the” include plural referents unless expressly limited to one or more referents otherwise.

[0021] 2. Numerical range: Unless otherwise expressly stated, all ranges or ratios disclosed herein will be understood to include any and all sub-ranges or sub-ratios contained therein. For example, a range or ratio of 1 to 30 stated should be considered to be included between a minimum value of 1 and a maximum value of 30, and any sub-range or sub-ratio, integer, decimal, or sub-range or sub-ratio consisting of integers or decimals including the end points.

[0022] The following non-limiting examples can enable those skilled in the art to more fully understand the present invention, but do not limit the present invention in any way. The following content is merely an exemplary description of the scope of the present invention, and those skilled in the art can make various changes and modifications to the present invention based on the disclosed content, which should also fall within the scope of the present invention.

[0023] The present invention is further described below by way of specific examples. The various chemical reagents used in the examples of the present invention are obtained through conventional commercial channels unless otherwise specified. Unless otherwise specified, the contents described below are all weight contents. Unless otherwise specified, it is understood that the process is carried out at room temperature.

[0024] In the following examples, some of the reagents are from the following Table 1: Table 1

[0025] In the following examples, the instruments and equipment used are shown in Table 2: Table 2

[0026] Example 1 A method for rapid thin layer analysis to determine the content of artemisinin in Artemisia annua, the steps are as follows: S1. Grind the Artemisia annua medicinal material to be tested, pass it through an 80-mesh sieve, mix it with ethyl acetate, perform ultrasonic extraction for 45 minutes, and filter it to obtain a test solution; the ratio of the mass of the Artemisia annua medicinal material to the volume of ethyl acetate is 1 g: 3 mL; S2. Take 0.1 μL of the test solution obtained in step S1 and spot it on the silica gel G thin layer plate to obtain the spotted silica gel G thin layer plate. Petroleum ether and ethyl acetate are mixed in a volume ratio of 5:1 to obtain a developing agent. Place the developing agent in a developing cylinder, seal the developing cylinder, and pre-saturate for 15 minutes. Place the spotted silica gel G thin layer plate into the developing cylinder, and the origin of the spot cannot be immersed in the developing agent. When the front edge of the developing agent goes up to the upper edge of the silica gel G thin layer plate and is 10% away from the origin of the spot, take out the silica gel G thin layer plate and blow dry it with a hair dryer to obtain a chromatography plate.

[0027] S3, coloring the chromatography plate obtained in step S2, spraying the vanillin sulfuric acid ethanol solution, heating at 105° C. for 10 minutes, cooling to room temperature, and obtaining a coloring chromatography plate.

[0028] The components of the vanillin sulfuric acid ethanol solution are, by mass percentage, 2.5% concentrated sulfuric acid, 15% vanillin, and the balance ethanol.

[0029] S4, compare the color chromatography plate obtained in step S3 with the standard colorimetric card. There are four color blocks on the standard colorimetric card: color block 1, color block 2, color block 3 and color block 4. The CMYK value of color block 1 is C=0, M=85, Y=70, K=0, which means that the content of artemisinin in Artemisia annua is 16-20 mg / g; the CMYK value of color block 2 is C=0, M=65, Y=50, K=0, which means that the content of artemisinin in Artemisia annua is 12-16 mg / g; the CMYK value of color block 3 is C=0, M=45, Y=30, K=0, which means that the content of artemisinin in Artemisia annua is 7-12 mg / g; the CMYK value of color block 4 is C=0, M=20, Y=10, K=0, which means that the content of artemisinin in Artemisia annua is 3-7 mg / g.

[0030] Example 2 Compared with Example 1, the difference is that in step S1, the ultrasonic extraction time is changed to 60 minutes, and the rest is the same.

[0031] Example 3 Compared with Example 1, the difference is that in step S1, the ultrasonic extraction time is changed to 30 minutes, and the rest is the same.

[0032] Example 4 Compared with Example 1, the difference is that in step S1, the ratio of the mass of Artemisia annua medicinal material to the volume of ethyl acetate is 1 g:4 mL, and the rest are the same.

[0033] Example 5 Compared with Example 1, the difference is that in step S1, the ratio of the mass of Artemisia annua medicinal material to the volume of ethyl acetate is 1 g:2 mL, and the rest are the same.

[0034] Example 6 Compared with Example 1, the difference is that in step S2, the developing solvent used is changed to a mixture of petroleum ether and ethyl acetate in a volume ratio of 15:1, and the rest is the same.

[0035] Example 7 Compared with Example 1, the difference is that in step S2, the developing solvent used is changed to a mixture of petroleum ether and ethyl acetate in a volume ratio of 10:1, and the rest is the same.

[0036] Example 8 Compared with Example 1, the difference is that in step S3, the mass composition of the sulfuric acid ethanol solution used is changed to 1% concentrated sulfuric acid, 20% vanillin, and the balance is ethanol, and the rest are the same.

[0037] Example 9 Compared with Example 1, the difference is that in step S3, the mass composition of the sulfuric acid ethanol solution used is changed to 5% concentrated sulfuric acid, 5% vanillin, and the balance is ethanol, and the rest are the same.

[0038] Comparative Example 1 Compared with Example 1, the difference is that in step S2, the developing agent is changed to a mixture of methanol and petroleum ether in a volume ratio of 1:5, and the rest is the same.

[0039] This method has the following problems in the process of determining the content of artemisia annua in artemisinin: During the development process, it seriously affects the measurement and judgment under ultraviolet light, especially when viewed under white light, the color development point of artemisinin on the silica gel G thin layer plate appears at the highest edge.

[0040] Comparative Example 2 Compared with Example 1, the difference is that in step S2, the developing agent is changed to a mixture of petroleum ether and ethyl acetate in a volume ratio of 0.2:1, and the rest is the same.

[0041] This method has the following problems in the process of determining the content of artemisia annua in artemisinin: During the development process, it seriously affects the measurement and judgment under ultraviolet light, especially under white light inspection. The color development point of artemisinin on the silica gel G thin layer plate appears at the highest edge.

[0042] Comparative Example 3 Compared with Example 1, the difference is that the mass percentage of concentrated sulfuric acid in the vanillin sulfuric acid ethanol solution is 10%, and the rest is the same.

[0043] This method has the following problems in the process of determining the content of artemisia annua in artemisinin: During the development process, it seriously affects the measurement and judgment under ultraviolet light, especially when inspected under white light, the color development point of artemisinin on the silica gel G thin layer plate tends to turn black and darker.

[0044] Comparative Example 4 Compared with Example 1, the difference is that in the vanillin sulfuric acid ethanol solution, the mass percentage content of the concentrated sulfuric acid is 0.1%, and the rest is the same.

[0045] This method has the following problems in the process of determining the content of artemisia annua in artemisinin: During the development process, the measurement and judgment under ultraviolet light are seriously affected, especially when inspected under white light, the color development point of artemisinin on the silica gel G thin layer plate is not clear.

[0046] Comparative Example 5 Compared with Example 1, the difference is that the colorimetric card used is different, and the rest is the same.

[0047] The colorimetric card used in Comparative Example 5 includes 4 color blocks: color block 1, color block 2, color block 3, and color block 4.

[0048] The CMYK values ​​of color block 1 are C=0, M=89, Y=95, K=0, which means that the content of artemisinin in Artemisia annua is 16-20 mg / g; the CMYK values ​​of color block 2 are C=0, M=55, Y=58, K=0, which means that the content of artemisinin in Artemisia annua is 12-16 mg / g; the CMYK values ​​of color block 3 are C=0, M=52, Y=44, K=0, which means that the content of artemisinin in Artemisia annua is 7-12 mg / g; the CMYK values ​​of color block 4 are C=0, M=20, Y=25, K=0, which means that the content of artemisinin in Artemisia annua is 3-7 mg / g.

[0049] Effect evaluation The content of artemisinin in Artemisia annua medicinal materials from different sources (Artemisia annua 1, Artemisia annua 2, Artemisia annua 3, Artemisia annua 4 and Artemisia annua 5) was detected. First, the content of artemisinin in Artemisia annua medicinal materials was determined by high performance liquid chromatography (General Rule 0512) recorded in Part II of the Chinese Pharmacopoeia. The experimental results are shown in Table 3: Table 3

[0050] The quantitative detection results are used as a basis for evaluating the accuracy of the detection methods provided in the embodiments and comparative examples of the present invention.

[0051] The evaluation process of the detection methods provided in each embodiment and comparative example was completed by 10 volunteers aged 25-50 years old. All volunteers had basic knowledge of analytical chemistry and were not color blind. Each volunteer tested the content of artemisinin in Artemisia annua medicinal materials from different sources according to the detection methods provided in each embodiment and comparative example, recorded the test results respectively, and compared them with the artemisinin content in Table 3 after summarizing. If the test result obtained by the pharmacopoeia detection method falls within the range of the sample test result obtained by the tester (such as: the artemisinin content in the Artemisia annua sample detected by the pharmacopoeia HPLC method is 15.89 mg / g, and the tester detects the artemisinin content according to the embodiment method as 12-16 mg / g, and the former falls within the range of the latter), it is regarded as "the one who obtains the correct test result". Calculate the accuracy rate between operators.

[0052] Inter-operator accuracy rate = number of people who obtained correct test results ÷ total number of people tested × 100%.

[0053] The results of the largest number of people who obtained the analysis results in each embodiment and comparative example were taken as the final test results. If the test results obtained by the pharmacopoeia test method fall within the range of the final test results, it is considered to be "correct results obtained". The correct rate between samples was calculated.

[0054] The accuracy rate between samples = the number of samples with correct results ÷ the total number of samples × 100%.

[0055] In particular, if the method has multiple analysis results with the largest number of people, it means that the test results are scattered and it is deemed that the correct result cannot be determined (for example, the content of artemisinin in the Artemisia annua sample detected by the pharmacopoeia HPLC method is 15.89 mg / g. Among the 10 testers, 5 testers respectively detected the artemisinin content as 12-16 mg / g and 7-12 mg / g according to the method of the embodiment, which means that the test results are scattered and the correct result cannot be obtained).

[0056] The volunteers used the detection methods of the embodiments and comparative examples to detect the content of artemisinin in Artemisia annua 1. The experimental results are summarized in Table 4: Table 4

[0057] Volunteers used the detection methods of various embodiments and comparative examples to detect the content of artemisinin in Artemisia annua 2. The experimental results are summarized in Table 5: Table 5

[0058] The volunteers used the detection methods of the embodiments and comparative examples to detect the content of artemisinin in Artemisia annua 3. The experimental results are summarized in Table 6: Table 6

[0059] Volunteers used the detection methods of various embodiments and comparative examples to detect the content of artemisinin in Artemisia annua 4. The experimental results are summarized in Table 7: Table 7

[0060] Volunteers used the detection methods of various embodiments and comparative examples to detect the content of artemisinin in Artemisia annua 5. The experimental results are summarized in Table 8: Table 8

[0061] The experimental results in Tables 4 to 8 were further summarized, and the experimental results were compared with the artemisinin content detected by HPLC in Table 3 to calculate the correct rate between samples. The results are shown in Table 9.

[0062] Table 9

[0063] It can be seen that compared with the detection methods provided in the comparative examples, the detection method provided in the embodiment has higher inter-operator accuracy and inter-sample correctness, can achieve rapid semi-quantitative determination of the content of artemisinin in Artemisia annua, and effectively reduce the experimental error caused by visual colorimetry between different detectors.

[0064] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, rather than to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions of the technical solution of the present invention by ordinary technicians in this field do not deviate from the essence and scope of the technical solution of the present invention.

Claims

1. A method for determining the content of artemisinin in Artemisia annua by rapid thin layer analysis, characterized in that: The following steps are involved: S1. Mix the medicinal material to be tested with the solvent, perform ultrasonic extraction, and filter to obtain the test solution; S2, spotting the test solution obtained in step S1 on the thin layer plate, developing, and drying to obtain a chromatography plate; S3, using vanillin sulfuric acid ethanol solution to develop the chromatography plate obtained in step S2 to obtain a color-developing chromatography plate; S4, comparing the color chromatography plate obtained in step S3 with the standard colorimetric card to determine the value range of the artemisinin content; Wherein, in step S1, the solvent is ethyl acetate; the ultrasonic extraction time is 30-60 min; the ratio of the weight of the medicinal material to be tested to the volume of the solvent is 1:2-4, in units of g:mL; In step S2, the developing agent is a mixture of petroleum ether and ethyl acetate in a volume ratio of 5-15:1; In step S3, the vanillin sulfuric acid ethanol solution is prepared by mixing concentrated sulfuric acid, vanillin and ethanol, and the mass percentage of the concentrated sulfuric acid in the vanillin sulfuric acid ethanol solution is 1%-5%; the color development comprises the following steps: Spray the chromatographic plate obtained in step S2 with the vanillin sulfuric acid ethanol solution, and heat at 100-110° C. for 10-20 min; the mass composition of the vanillin sulfuric acid ethanol solution is: 1%-5% concentrated sulfuric acid, 5%-20% vanillin, and the balance is ethanol; The standard colorimetric card in step S4 includes color block 1, color block 2, color block 3 and color block 4; The CMYK values ​​of the color block 1 are C=0, M=85, Y=70, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 1 is 16-20 mg / g; The CMYK values ​​of the color block 2 are C=0, M=65, Y=50, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 2 is 12-16 mg / g; The CMYK values ​​of the color block 3 are C=0, M=45, Y=30, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 3 is 7-12 mg / g; The CMYK values ​​of the color block 4 are C=0, M=20, Y=10, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 4 is 3-7 mg / g.

2. The method according to claim 1, characterized in that In step S2, the developing agent is a mixture of petroleum ether and ethyl acetate in a volume ratio of 5:

1.

3. The method according to claim 1, characterized in that: In step S2, the developing agent is a mixture of petroleum ether and ethyl acetate in a volume ratio of 15:

1.

4. The method according to claim 1, characterized in that: In step S2, the thin layer plate is a silica gel G thin layer plate.

5. The method according to claim 1, characterized in that In step S2, the development is specifically as follows: placing the spotted silica gel G thin layer plate into a development cylinder, and the spot origin cannot be immersed in the developing agent; when the front edge of the developing agent rises to the upper edge of the silica gel G thin layer plate, 10%-30% away from the spot origin, the silica gel G thin layer plate is taken out.

6. The method according to claim 1, characterized in that In step S3, the mass composition of the vanillin sulfuric acid ethanol solution is: 5% concentrated sulfuric acid, 5% vanillin, and the balance is ethanol.

7. The method according to claim 1, characterized in that In step S3, the mass composition of the vanillin sulfuric acid ethanol solution is: 2.5% concentrated sulfuric acid, 15% vanillin, and the balance is ethanol.

8. The method according to claim 1, characterized in that In step S4, the comparative identification is specifically to place the sample at room temperature and then compare it with a standard colorimetric card under white light.

9. Use of the method according to any one of claims 1 to 8 in determining the content of artemisinin.

10. A standard colorimetric card, characterized in that: The standard colorimetric card includes color block 1, color block 2, color block 3 and color block 4; The CMYK values ​​of the color block 1 are C=0, M=85, Y=70, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 1 is 16-20 mg / g; The CMYK values ​​of the color block 2 are C=0, M=65, Y=50, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 2 is 12-16 mg / g; The CMYK values ​​of the color block 3 are C=0, M=45, Y=30, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 3 is 7-12 mg / g; The CMYK values ​​of the color block 4 are C=0, M=20, Y=10, K=0, and the content of artemisinin in the Artemisia annua represented by the color block 4 is 3-7 mg / g.

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