A mass analysis method for extracting effective components of gastrodia elata by hdes

Abstract

The application discloses a quality analysis method for extracting effective components of Gastrodia elata by using HDES, and comprises the following steps: (1) synthesizing HDES by using L-menthol and DL-lactic acid; (2) adding the HDES in step (1) into Gastrodia elata medicinal material powder, stirring to extract effective components, centrifuging and diluting the obtained mixture after the extraction is completed, obtaining a test sample solution, and performing chromatographic analysis to determine the content of the effective components; (3) detecting the quality indexes of the test sample solution in step (2); and (4) analyzing the quality of Gastrodia elata by using a chemometrics analysis method. The extraction method is green, simple, fast and high in extraction efficiency, and is favorable for the extraction of effective components in medicine and food homologous medicinal materials.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of chemical extraction analysis, and particularly relates to a quality analysis method for extracting effective components of Gastrodia elata Bl. by using HDES. BACKGROUND

[0002] Gastrodia elata Bl. is the dried tuber of Gastrodia elata Bl. of Orchidaceae, which is used as a medicinal material and is a functional food with high nutritional value and medicinal value. It is one of the “ten famous medicinal materials” in Yunnan and is regarded as the “top product” in traditional Chinese medicine. It was included in the list of medicinal and edible materials in 2019. As a medicinal and dietary source, Gastrodia elata Bl. mainly contains active substances such as aromatic compounds, phenols, organic acid esters, sugars and their glycosides, and sterols. Studies have shown that Gastrodia elata Bl. has various medicinal effects and is commonly used in clinical practice to treat ischemic dementia, ischemic myocardial injury, hypertension, atherosclerosis, and cerebrovascular dizziness. It also has certain effects on Parkinson's disease, aging, and migraine and other nervous system diseases. Among the numerous active substances in Gastrodia elata Bl., phenolic compounds are increasingly attracting the attention of researchers and the industry due to their potential effects of promoting blood circulation and relieving nerves.

[0003] The effective components of medicinal and edible materials are affected by different production areas, processing methods, and extraction methods, which in turn affects their medicinal effects. The extraction of effective components of most medicinal and edible materials still uses traditional organic solvents such as methanol, ethanol, diethyl ether, ethyl acetate, chloroform, and petroleum ether. However, these traditional solvents have the disadvantages of long extraction time, large solvent usage, low extraction efficiency, easy evaporation, high toxicity, and non-biodegradability. In addition, the long-term use of traditional solvents can pose a threat to human health and harm the ecological environment. Therefore, it is crucial to develop a new green solvent with high extraction efficiency, low volatility, and stable properties to replace traditional organic solvents.

[0004] In addition, the medicinal effects of medicinal and edible materials are often the result of the combined action of multiple components, and the content of a single component often changes due to different storage conditions and processes. Therefore, monitoring multiple components rather than a single component can reflect the relative stability and comprehensiveness of the quality of medicinal materials. Chemometrics analysis has the advantages of efficient data processing, accurate information extraction, and flexible methods, and plays an important role in environmental analysis, food science, drug development, and quality control. Therefore, a new method for efficiently extracting three main phenolic compounds in Gastrodia elata Bl. is established, and combined with chemometric analysis, the evaluation results are more comprehensive and reasonable, which can provide a more scientific theoretical and data basis for the quality control and quality improvement of medicinal materials. SUMMARY

[0005] The application aims to provide a quality analysis method for extracting effective components of Gastrodia elata by using HDES, which has the advantages of green, simple, fast and high extraction efficiency.

[0006] To achieve the above-mentioned application purposes, the application adopts the following technical solutions:

[0007] The application provides a quality analysis method for extracting effective components of Gastrodia elata by using HDES, which comprises the following steps:

[0008] (1) L-menthol and DL-lactic acid are mixed in a molar ratio of 1:1-7, and then the mixture is stirred in an oil bath at 60°C for 30 min to obtain a uniform transparent liquid as HDES;

[0009] (2) The HDES in step (1) is added to Gastrodia elata medicinal material powder, and the effective components are extracted by stirring, and after the extraction is completed, the obtained mixture is centrifuged and diluted to obtain a test solution, and chromatographic analysis is performed to determine the content of the effective components;

[0010] (3) The quality indicators of the test solution in step (2) are detected;

[0011] (4) Under the optimal conditions explored, the same sample is injected 6 times to verify the method precision; the stability of the sample solution is evaluated by measuring the test solution after being placed at room temperature for 0, 2, 4, 8, 16 and 24 h; the repeatability of the method is evaluated by measuring 6 test solutions; in addition, standard solutions are added to samples with known contents of three phenolic compounds in Gastrodia elata to determine the sample recovery rate; the corresponding spectral data obtained are analyzed;

[0012] (5) Combined with the quality indicators in step (3), the quality of Gastrodia elata is comprehensively evaluated by chemometrics analysis.

[0013] Preferably, the molar ratio of L-menthol to DL-lactic acid in step (1) is 1:4.

[0014] Further, the solid-liquid ratio (g / mL) of Gastrodia elata medicinal material powder to HDES in step (2) is 1:6-1:36.

[0015] Preferably, the solid-liquid ratio (g / mL) of Gastrodia elata medicinal material powder to HDES in step (2) is 1:24.

[0016] Further, the stirring extraction conditions in step (2) are as follows: the stirring extraction speed is 1000 rpm / min, and the stirring extraction time is 10-60 min.

[0017] Preferably, the stirring extraction time in step (2) is 20 min.

[0018] Further, the chromatography in step (2) is HPLC, and the conditions are as follows: the chromatographic column is ZORBAX Extend-C 18 ; the mobile phase is acetonitrile C and 0.1% phosphoric acid solution D; the gradient elution sequence is 0-20 min, 5% C, 95% D; 20-30 min, 5%-15% C, 95%-85% D; 30-40 min, 15%-20% C, 85%-80% D; 40-55 min, 20% C, 80% D; 55-60 min, 20%-5% C, 80%-95% D; the flow rate is 1.0 mL / min; the detection wavelength is 280 nm; the column temperature is 25°C, and the injection volume is 10 µL.

[0019] Further, the mixture in step (2) is diluted with methanol.

[0020] Further, the effective components are gastrodin, p-hydroxybenzyl alcohol and balichensin A.

[0021] Further, the quality indicators in step (3) include moisture, total ash, alcohol-soluble extract and metal element content.

[0022] Further, the chemometric analysis in step (5) includes Pearson correlation analysis, cluster analysis, principal component analysis and factor analysis.

[0023] Further, when the chemometric analysis in step (5) is performed, the Cu element has significant correlation with the effective components.

[0024] Further, in step (4), the RSD of the precision of the content of the three phenolic compounds in the gastrodin sample is 0.47-0.96%, the RSD of the repeatability is 0.98-1.52%, the RSD of the solution stability is 0.61-1.33%, and the sample recovery rate is 100.35-112.10%.

[0025] Compared with the prior art, the present application has the following advantages and beneficial effects:

[0026] 1. The present application first uses a hydrophobic deep eutectic solvent (HDES) to efficiently extract phenolic compounds in gastrodin, i.e., gastrodin, p-hydroxybenzyl alcohol and balichensin A.

[0027] 2. The present application establishes a green, simple and rapid new method for extracting effective components of a homophylous medicinal material and performing quality analysis based on chemometric analysis.

[0028] 3、Compared with the traditional extraction method, the extraction rates of gastrodin, p-hydroxybenzyl alcohol and bartzin A are 2.8, 2.5 and 16.1 times higher than those of methanol extraction, respectively, so the extraction efficiency of the method is significantly improved.

[0029] 4、The present application verifies that the content of gastrodin has significant correlation with the content of Cu element. BRIEF DESCRIPTION OF DRAWINGS

[0030] Figure 1 The infrared spectrum of L-menthol, DL-lactic acid and HDES.

[0031] Figure 2 The single factor optimization diagram of extracting three phenolic compounds of gastrodin by HDES, wherein, figure A is the extraction method optimization diagram; figure B is the kind of HDES optimization diagram; figure C is the extraction time optimization diagram; figure D is the solid-liquid ratio optimization diagram; figure E is the molar ratio optimization diagram of HDES.

[0032] Figure 3 The determination result diagram of moisture, total ash and alcohol-soluble extract of 15 batches of gastrodin samples.

[0033] Figure 4 The correlation heat map of phenolic compounds, moisture, total ash, alcohol-soluble extract and metal element content in gastrodin.

[0034] Figure 5 The average linkage (intergroup) phylogenetic diagram of 15 batches of gastrodin, wherein figure A takes the content of gastrodin, p-hydroxybenzyl alcohol, bartzin A, moisture, total ash, alcohol-soluble extract and 7 kinds of metal elements as a data matrix; figure B takes the content of gastrodin, p-hydroxybenzyl alcohol, bartzin A, moisture, total ash, alcohol-soluble extract and Cu element as a data matrix; figure C takes the content of gastrodin, p-hydroxybenzyl alcohol, bartzin A, moisture, total ash and alcohol-soluble extract as a data matrix.

[0035] Figure 6 The principal component analysis diagram of 15 batches of gastrodin, wherein figure A carries out principal component analysis on the content of gastrodin, p-hydroxybenzyl alcohol, bartzin A, moisture, total ash, alcohol-soluble extract and 7 kinds of metal elements; figure B carries out principal component analysis on the content of gastrodin, p-hydroxybenzyl alcohol, bartzin A, moisture, total ash, alcohol-soluble extract and Cu element; figure C carries out principal component analysis on the content of gastrodin, p-hydroxybenzyl alcohol, bartzin A, moisture, total ash and alcohol-soluble extract. DETAILED DESCRIPTION

[0036] The technical solutions of the present application are further described in combination with the following specific examples.

[0037] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods, and the materials and reagents used can be purchased from biological or chemical reagent companies.

[0038] Example 1

[0039] I. Preparation and Characterization of Hydrophobic Deep Eutectic Solvent (HDES)

[0040] Using L-menthol as HBA and DL-lactic acid as HBD, HBA and HBD were mixed in a round-bottom flask at a molar ratio of 1:4. The mixture was then stirred in an oil bath at 60°C for 30 min to obtain a homogeneous and transparent liquid, HDES.

[0041] To confirm the formation of hydrogen bonds, the FT-IR spectra of pure L-menthol, DL-lactic acid, and HDES were compared, such as... Figure 1 As shown. The hydroxyl stretching band (O–H) of L-menthol extends from 3261 cm. −1 Blue shifted to 3407 cm −1 DL-lactic acid at 1720 cm −1 The characteristic spectral band at this location is the C=O stretching vibration, which starts from 1720 cm⁻¹. −1 Redshifted to 1717 cm −1 This demonstrates that intermolecular hydrogen bonds formed between L-menthol and DL-lactic acid, indicating successful preparation of the HDES. Characteristic peaks of both L-menthol and DL-lactic acid are shown in the HDES spectrum. Furthermore, comparing the FT-IR spectrum of the HDES with that of its components revealed no new peak disappearances or appearances, indicating that no reaction occurred during the HDES preparation process.

[0042] II. Extraction of effective components from Gastrodia elata

[0043] Accurately weigh 0.5000 g of Gastrodia elata powder and add 12 mL of the synthesized HDES. Stir at 1000 rpm / min for 20 min at room temperature. After extraction, centrifuge the mixture at 8000 rpm for 5 min. Dilute the final sample 3 times with methanol to reduce viscosity and prepare the test solution. Finally, perform HPLC analysis to extract the active ingredients gastrodin, p-hydroxybenzyl alcohol, and baicalin A.

[0044] HPLC conditions: Column: ZORBAX Extend-C 18(4.6 mm´250 mm, 5 µm). The mobile phase was acetonitrile (C) and 0.1% phosphoric acid solution (D); the gradient elution sequence was: 0−20 min, 5% C, 95% D; 20−30 min, 5%−15% C, 95%−85% D; 30−40 min, 15%−20% C, 85%−80% D; 40−55 min, 20% C, 80% D; 55−60 min, 20%−5% C, 80%−95% D; the flow rate was 1.0 mL / min; the detection wavelength was 280 nm; the column temperature was 25 °C, and the injection volume was 10 µL.

[0045] An appropriate amount of gastrodin, p-hydroxybenzyl alcohol and balichensin A was precisely weighed, dissolved in methanol to prepare a mixed reference solution and a reference solution for determination.

[0046] The established method was verified for linear relationship and range, precision, solution stability, repeatability and spiked recovery rate. For example, the same sample was injected 6 times to verify the method precision; the stability of the sample solution was evaluated by measuring the test sample solution after being placed at room temperature for 0, 2, 4, 8, 16 and 24 h; the repeatability of the method was evaluated by measuring 6 test sample solutions; in addition, the standard solution was added to the sample with known content of the three phenolic compounds to determine the sample recovery rate.

[0047] The RSD of the precision of the content of the three phenolic compounds in the Gastrodia rubra sample was 0.47−0.96%, the RSD of the repeatability was 0.98−1.52%, the RSD of the solution stability was 0.61−1.33%, and the recovery rate was 100.35−112.10%.

[0048] III. Single factor investigation

[0049] 1. Extraction method

[0050] Common effective ingredient extraction methods include ultrasonic extraction, stirring extraction, microwave extraction, cold soaking extraction, reflux extraction and Soxhlet extraction. Among them, ultrasonic extraction and stirring extraction have the advantages of low energy consumption, simple operation, high extraction efficiency and wide application, so this study mainly compares these two extraction methods. 0.5000 g of Gastrodia rubra medicinal material (S2) was precisely weighed in a centrifuge tube, and HDES (L-menthol: DL-lactic acid) was used as the extraction solvent. The mixture was ultrasonically extracted (power 500 W) and stirred at room temperature for 60 min to extract the three components (gastrodin, p-hydroxybenzyl alcohol and balichensin A). After extraction, the mixture was centrifuged at 8000 rpm for 5 min. The final sample was diluted 3 times with methanol to reduce the viscosity, and then analyzed by HPLC.

[0051] The results are shown in Table 1.Figure 2 A shows that the extraction amount is larger using stirring extraction. Therefore, the subsequent experiment uses stirring extraction.

[0052] 2. Selection of HDES types

[0053] Different HDESs significantly affect the polarity and hydrogen bond network of HDESs due to their different components, thereby affecting their solubility. Therefore, different HDESs can extract different polar effective components. Taking Gastrodia elata medicinal materials, HDES-1 (L-menthol: n-hexanoic acid, 1:1), HDES-2 (L-menthol: nonanoic acid, 1:1), HDES-3 (L-menthol: n-decanoic acid, 1:1), HDES-4 (L-menthol: n-octanol, 1:1), and HDES-5 (L-menthol: DL-lactic acid, 1:1) were used as extraction solvents to extract three phenolic compounds, obtain test sample solutions, and finally perform HPLC analysis to calculate the extraction amounts of the three components.

[0054] The results are shown in Figure 2 B shows that using HDES-5 (L-menthol: DL-lactic acid, 1:1) as the extraction solvent, the extraction amount is the largest. Therefore, the subsequent content determination experiment uses HDES-5 for extraction.

[0055] 3. Effect of extraction time

[0056] Extraction time has an important influence on the extraction amount of effective components. Generally speaking, too short extraction time may result in incomplete dissolution or transfer of effective components to the solution, and low extraction efficiency; while too long extraction time will result in unnecessary energy waste, and may introduce other components. Taking Gastrodia elata medicinal materials, stirring extraction was performed for different times of 10-60 min to obtain test sample solutions.

[0057] The results are shown in Figure 2 C shows that after stirring for more than 20 min, the extraction amounts of the components tend to be flat, and the optimal extraction time is selected as 20 min in the principle of simple operation and less time consumption.

[0058] 4. Effect of solid-liquid ratio

[0059] Too high or too low solid-liquid ratio may affect the extraction efficiency and purity of the extracted substances. Too high solid-liquid ratio may result in solvent waste, low extraction efficiency, or introduction of impurities; too low solid-liquid ratio may result in insufficient coverage of the raw materials by the solvent, thereby affecting the extraction effect. Taking Gastrodia elata medicinal materials, HDES was added at different solid-liquid ratios (g / mL) of 1:6-1:36 to obtain test sample solutions and perform determination.

[0060] The results are shown in Figure 2D shows that the extraction rate gradually increases when the solid-liquid ratio (g / mL) is 1 :6-1 :24. The extraction amount of each index component changes little with the increase of liquid amount. Insufficient liquid amount will lead to incomplete extraction, while excessive liquid amount will cause waste of extractant. Therefore, the solid-liquid ratio (g / mL) of 1 :24 is selected for subsequent experiments.

[0061] 5. Effect of HDES molar ratio

[0062] When the molar ratio of HDES is different, the structural stability, viscosity, polarity and melting point of HDES will also be different. An appropriate amount of Gastrodia elata medicinal materials is selected, and 1 : 1-7, 7 different HDES molar ratios are selected to extract three phenolic compounds to obtain the test solution.

[0063] The results are shown in Figure 2 E shows that when the molar ratio of L-menthol and DL-lactic acid is 1 :4, the extraction amount of the three index components is the largest.

[0064] Example 2

[0065] 1. Comparison of extraction efficiency of HDES and traditional methanol

[0066] In order to compare the extraction effects of HDES and methanol on three phenolic compounds in Gastrodia elata, Gastrodia elata medicinal material powder is prepared according to the steps of Example 1 to obtain the test solution, which is determined by HPLC. Methanol extraction is carried out according to the method reported in the literature, that is, 2.0000 g of Gastrodia elata medicinal material powder is accurately weighed in a 50 mL glass conical flask with a plug, 25 mL of 60% methanol is added, the mass is determined, ultrasonic is performed for 60 min, after cooling, the lost mass is made up with 60% methanol, shaken, filtered with a microporous filter membrane (0.45 μm), and the test solution is obtained and determined. The corresponding extraction rate is calculated, and the extraction rates of the two extraction solvents are compared.

[0067] The results are shown in Table 1. According to the results in the table, the extraction rates of HDES for gastrodin, p-hydroxybenzyl alcohol and balichensin glycoside A are 2.8, 2.5 and 16.1 times higher than those of methanol extraction, respectively. It can be seen that the extraction rate of HDES for three main phenolic compounds in Gastrodia elata is greatly improved compared with traditional methanol extraction, especially for balichensin glycoside A.

[0068] Table 1 Comparison of extraction rates of HDES and methanol

[0069]

[0070] 2. Determination of active ingredient content

[0071] Accurately weigh 0.5000 g of Gastrodia elata powder from 15 different origins (see Table 2 for details) to prepare test solutions, which were then injected for analysis. The contents of gastrodin, p-hydroxybenzyl alcohol, and barisonin A were calculated using the external standard method.

[0072] Table 2. Numbering of Gastrodia elata samples from different origins

[0073]

[0074] The results are shown in Table 3. The contents of gastrodin, p-hydroxybenzyl alcohol, and barisonoside A were 3.4–18.8 mg / g, 0.8–3.4 mg / g, and 49.6–167.8 mg / g, respectively, indicating that the contents of the indicative components of Gastrodia elata varied from different origins. Regarding the extraction yield of the three phenolic compounds by HDES, barisonoside A was extracted in the highest amount, while p-hydroxybenzyl alcohol was extracted in the lowest amount. Specifically, HDES extraction of S14 yielded the highest content of p-hydroxybenzyl alcohol, while S9 yielded the highest contents of gastrodin and barisonoside A. Overall, the contents of the three indicative components were relatively high in S6, S9, and S12.

[0075] Table 3. Results of determination of effective components in Gastrodia elata from different origins

[0076]

[0077] 3. Determination of moisture, total ash and alcohol-soluble extract.

[0078] The determination of moisture, total ash, and alcohol-soluble extracts is of great significance for accurately evaluating the quality of medicinal and edible herbs and ensuring their stable efficacy during use. Appropriate amounts of Gastrodia elata powder from 15 different origins were taken, and the contents of moisture, total ash, and alcohol-soluble extracts were determined according to the corresponding methods in the General Chapters, Part IV of the 2020 edition of the Chinese Pharmacopoeia.

[0079] The results are as follows Figure 3 As shown, the moisture content of all 15 batches of Gastrodia elata samples was less than 15.0%, the total ash content was less than 4.5%, and the alcohol-soluble extract content was greater than 15.0%, indicating that the moisture, total ash, and alcohol-soluble extract content of the Gastrodia elata samples from 15 different producing areas were all qualified.

[0080] 4. Determination of the content of metal elements in Gastrodia elata medicinal material

[0081] During the planting and processing of Chinese herbal medicines, heavy metal pollution may occur due to non-standard planting, misuse of various pesticide formulations, or immature technology, which may affect the quality and safety of Chinese herbal medicines. Therefore, the analysis and determination of metal elements are also an important part of evaluating the quality of Chinese herbal medicines. In this study, the contents of seven metal elements in Gastrodia elata from 15 different origins were determined by ICP-OES.

[0082] The results are shown in Table 4. The metal element content determination results of Gastrodia elata from different origins were different, and the content of some metal elements differed greatly. This also verified that the content of metal elements was closely related to the origin.

[0083] Table 4 Determination results of metal elements

[0084]

[0085] ND: not detected

[0086] Example 3: Chemometric analysis

[0087] 1. Pearson correlation analysis

[0088] Pearson correlation analysis was performed between the "quality indicators" of Gastrodia elata, such as the contents of three phenolic compounds, moisture, total ash, alcohol-soluble extract, and the contents of seven metal elements.

[0089] The results are shown in Figure 4 As shown in the figure, the content of three phenolic compounds and the content of the remaining ten variables had significant differences in the content of gastrodin and Cu element. The contents of the other two components had no significant correlation with the contents of the remaining variables. The total ash content had significant differences with the contents of Cd and Pb metal elements. The contents of the seven metal elements mostly had certain correlation with each other.

[0090] 2. Cluster analysis

[0091] Using IBM SPSS Statistics 23 software, the contents of "quality indicators" and seven metal elements were used as data matrix, the between-groups linkage method was used, the squared Euclidean distance was selected as the scale, and the phylogenetic diagram was used to represent the systematic cluster analysis of Gastrodia elata samples from 15 origins. The phylogenetic diagram is shown in Figure 5A, can intuitively reflect the classification of different samples of Gastrodia elata in this study. Cluster analysis results show that when the square Euclidean distance is 5, 15 batches of Gastrodia elata samples can be obviously clustered into 5 groups. The first group (3 producing areas): S2, S11, S15; the second group (3 producing areas): S1, S4, S7; the third group (5 producing areas): S9, S12, S6, S13, S14; the fourth group (3 producing areas): S5, S10, S3; the fifth group (1 producing area): S8. However, from the above correlation analysis results, among the many variables, only the content of gastrodin and the content of Cu element have significant correlation. Therefore, in order to verify the correlation analysis results, the content of "quality indicators" and the content of Cu element were used as data matrices, and the content of "quality indicators" was used as data matrix, respectively, for system clustering analysis. The clustering results of the two are the same, see Figure 5 B and 5C. Among them, S2, S3, S1 in the first group of samples and S11, S5 are each clustered into a small class, S4, S5, S7, S10, S13 and S8, S14, S6 in the second group of samples are each clustered into a small class, and the samples show certain intra-group differences; the third group of samples is clustered more concentratedly, and the sample quality consistency is better.

[0092] In summary, compared with the contents of other metal elements, the content of Cu element has the same distribution rule as the contents of three phenolic compounds, moisture, total ash and alcohol-soluble extract, which closely affect the quality of Gastrodia elata, and the clustering analysis results are more consistent with the Pearson correlation analysis results.

[0093] 3. Principal component analysis

[0094] The contents of "quality indicators" and seven metal elements in 15 producing areas of Gastrodia elata samples were subjected to principal component analysis. According to the analysis, the Kaiser-Meyer-Olkin (KMO) and Bartlett's sphericity test results were KMO=0.359, P=0.000, which did not meet the KMO>0.6 condition of the application program data suitability requirement of principal component analysis, and in addition, from the correlation matrix of the data matrix, it can be seen that the correlation between the variables is not significant, and the correlation coefficient is less than 0.5. Therefore, the data matrix does not meet the data suitability requirement of principal component analysis, and the principal component analysis cannot be carried out. Figure 6As shown in Figure A, the samples are relatively dispersed, and the classification results differ significantly from the cluster analysis results, indicating that the data corresponding to the above variables are not suitable for principal component analysis. Based on the results of cluster analysis and Pearson correlation analysis, principal component analysis was performed on the data of "quality index" content and Cu element content. The KMO value and Bartlett's test of sphericity showed that KMO > 0.6, P < 0.05, indicating significant data correlation, making it suitable for principal component analysis and factor analysis. In the analysis results of the two sets of different variable data, the two principal components (PC1 and PC2) with eigenvalues ​​greater than 1 were selected, with cumulative contribution rates of 63.75% and 70.05%, respectively. This indicates that the first two principal components can represent 63.75% of the information of the seven components of Gastrodia elata and 70.05% of the information of the six components of Gastrodia elata, respectively. Therefore, the two principal components can basically reflect the information of the measured corresponding components. Figure 6 As shown in B and 6C, gastrodin, p-hydroxybenzyl alcohol, barisoniside A, alcohol-soluble extract, and total ash showed a strong positive correlation with PC1, while moisture and Cu content showed a strong positive correlation with PC2. In addition, alcohol-soluble extract also showed a strong correlation with p-hydroxybenzyl alcohol. The 15 batches of samples were divided into 3 categories, and the results were basically consistent with the cluster analysis results, which further verified the classification results of the cluster analysis.

[0095] 4. Factor analysis

[0096] In factor analysis, a higher overall score indicates better product quality. Factor analysis was performed on the content of "quality indicators" and Cu element content. The eigenvectors were multiplied by the standardized data to obtain two principal component scores (F1 and F2). The weights were calculated as follows: the ratio of the eigenvalue corresponding to each principal component to the sum of the total eigenvalues ​​of the extracted principal components. The principal component comprehensive model was obtained. The overall score (F1) of 15 batches of Gastrodia elata samples was calculated according to formula (2). j (See Tables 5 and 6). The calculation results show that the principal component comprehensive scores of the samples from the 15 producing areas were -1.043 to 1.352 and -1.029 to 1.353, respectively, under both conditions. Ranking the comprehensive scores under both conditions yielded the same results: S9, S12, and S6 ranked in the top three, indicating that the quality of *Gastrodia elata* from these three producing areas was relatively superior. This result is consistent with the determination results of the indicator component content. Therefore, factor analysis can be used to evaluate the quality differences of *Gastrodia elata* samples from different producing areas.

[0097] (1)

[0098] (2)

[0099] Among them, W tFtis the relative weight of the tth principal component, C t Ftis the variance contribution rate of the tth principal component, C z Ftis the cumulative variance contribution rate. F1and F2are the corresponding principal component scores, F j Ftis the comprehensive score.

[0100] Table 5 Principal component scores and ranking of 15 batches of Gastrodia elata samples

[0101]

[0102] Data matrix analysis was performed on the phenolic compounds, moisture, total ash, alcohol-soluble extract, and Cu content of 15 batches of GE samples.

[0103] Table 6 Principal component scores and ranking of 15 batches of Gastrodia elata samples

[0104]

[0105] Data matrix analysis was performed on the phenolic compounds, moisture, total ash, and alcohol-soluble extract content of 15 batches of GE samples.

[0106] The above examples are only used to illustrate the technical solutions of the present application, but not to limit it; although the present application has been described in detail with reference to the foregoing examples, the technical solutions recorded in the foregoing examples can still be modified by those of ordinary skill in the art, or some technical features thereof can be replaced by equivalents; and these modifications or replacements do not make the corresponding technical solutions deviate from the spirit and scope of the technical solutions claimed by the present application.

Claims

1. A quality analysis method for extracting effective components from Gastrodia elata using the hydrophobic deep eutectic solvent HDES, characterized in that, Includes the following steps: (1) Mix L-menthol and DL-lactic acid in a molar ratio of 1:1–7, and then stir the mixture in an oil bath at 60°C to obtain a uniform and transparent liquid called HDES. (2) Add the HDES from step (1) to the Gastrodia elata powder, with a material-to-liquid ratio of Gastrodia elata powder to HDES of 1:6-1:36; stir to extract the active ingredients. After extraction, centrifuge and dilute the mixture to obtain the test solution, and perform chromatographic analysis to determine the content of active ingredients. The chromatography method was HPLC, and the conditions were as follows: HPLC column: ZORBAX Extend-C 18 The mobile phase was acetonitrile (C) and 0.1% phosphoric acid solution (D); the gradient elution sequence was: 0–20 min, 5% C, 95% D; 20–30 min, 5%–15% C, 95%–85% D; 30–40 min, 15%–20% C, 85%–80% D. 40–55 min, 20% C, 80% D; 55–60 min, 20%–5% C, 80%–95% D; flow rate 1.0 mL / min; detection wavelength 280 nm; column temperature 25℃; injection volume 10 µL. The conditions for the stirring extraction are: stirring extraction speed of 1000 rpm / min and stirring extraction time of 10–60 min; the active ingredients are gastrodin, p-hydroxybenzyl alcohol and barisonin A; (3) Detect the quality indicators of the test solution in step (2); (4) Combine the quality indicators of step (3) and use chemometric analysis to analyze the quality of Gastrodia elata.

2. The method according to claim 1, characterized in that, In step (2), the mixture is diluted with methanol.

3. The method according to claim 1, characterized in that, The quality indicators in step (3) include moisture, total ash, alcohol-soluble extract and metal element content.

4. The method according to claim 1, characterized in that, The chemometric analysis in step (4) includes Pearson correlation analysis, cluster analysis, principal component analysis and factor analysis.

5. The method according to claim 1, characterized in that, When performing chemometric analysis in step (4), there is a significant correlation between Cu element and effective components.

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