CN111521724A - Method for screening anti-fatigue components of health wine - Google Patents
Method for screening anti-fatigue components of health wine Download PDFInfo
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Abstract
The invention discloses a method for screening anti-fatigue components of health-care wine, which comprises the following steps: carrying out qualitative analysis on the components of the health wine; carrying out quantitative analysis on characteristic components of the health-care wine; evaluating the anti-fatigue efficacy by using a zebra fish model; and screening the anti-fatigue main effect component group by using a least square method. The invention has the following advantages: (1) comprehensive qualitative analysis is carried out on the components of the health-care wine by adopting HPLC-QTOF-MS/MS and GC-MS technologies; (2) quantitative analysis is carried out on the monitorable components of the health-care wine by adopting an HPLC-QQQ-MS quantitative analysis method and a GC-MS quantitative analysis method; (3) the anti-fatigue effect of the health-care wine is evaluated through the zebra fish model, and the anti-fatigue main active ingredient group is screened through the least square method, so that the quality control of the anti-fatigue ingredients of the health-care wine is realized.
Description
Technical Field
The invention relates to the field of quality control of prepared wine, and particularly relates to a screening method of anti-fatigue components of health-care wine.
Background
The quality control method is a quality management method for ensuring the product quality and improving the product quality continuously. According to the method, the distribution of product quality data is researched and analyzed, the law of quality difference is revealed, the reasons influencing the quality difference are found out, technical organization measures are adopted, and factors generating inferior-quality products or unqualified products are eliminated or controlled, so that each link of the product in the whole production process can be normally and ideally carried out, and finally the product can reach the natural attributes and characteristics required by people, namely the applicability, reliability and economy of the product.
The health wine is prepared from Chinese medicinal materials with nourishing and treating functions and wine, belongs to one kind of compound wine, and has the functions of preventing and treating diseases and strengthening body. In the quality control of health wine, different detection means are adopted to ensure the stability of various component indexes of the wine. Patent CN1991361 discloses a method for detecting the quality of chinese health wine by using a fingerprint technology, which determines the quality of health wine by establishing a database of chinese health wine fingerprint spectra and comparing and analyzing the samples during detection.
Disclosure of Invention
The invention relates to a method for screening anti-fatigue components of health-care wine, which establishes a method for controlling the quality of the anti-fatigue components of the health-care wine by qualitatively and quantitatively analyzing the components of the health-care wine, evaluating the anti-fatigue drug effect of a zebra fish model and screening anti-fatigue main active component groups.
The scheme of the invention is as follows:
(1) qualitative analysis of health wine components
The HPLC-QTOF-MS/MS technology is adopted to carry out systematic identification research on chemical components contained in the health-care wine.
Chromatographic conditions are as follows: the column temperature is 25 ℃; mobile phase: water (containing formic acid with certain concentration) (A) -acetonitrile (containing formic acid with certain concentration) (B); flow rate: 0.8 mL/min; sample introduction volume: 2 μ L. The gradient elution conditions were: 0-2.5min, 10% B; 2.5-20min, 10-20% B; 20-35min, 20-35% B; 35-40min, 35-60% B; 40-45min, 60-90% B; 45-50min, 90-95% B; 50-60min, 95% B.
Mass spectrum conditions: flow rate of dry nitrogen: 10L/min; temperature of dry nitrogen: 350 ℃; atomization pressure: 35 psig; temperature of sheath gas: 350 ℃; flow rate of sheath gas: 11.0L/min; capillary voltage: 4000V; taper hole voltage: 65V; OCT 1RF Vpp: 750V; cleavage voltage: 135V; secondary collision energy: 15V, 25V, 35V and 50V. And (3) monitoring in a full scanning mode in a positive ion mode and a negative ion mode, wherein the scanning range of the mass number of the ions is set to be m/z 50-1700.
The GC-MS technology is adopted to systematically identify volatile components from angelica, fructus amomi, rhizoma curculiginis, fructus alpiniae oxyphyllae, cinnamon and clove in the health care wine.
Chromatographic conditions are as follows: the instrument comprises the following steps: agilent 7890B GC-5977A MSD; a chromatographic column: agilent19091S-433UI HP-5ms Ultra Inert (30 m. times.250 μm,0.25 μm); 0-325 ℃; carrier gas: 99.999% high-purity He; flow rate: 1 mL/min; sample introduction volume: 1 μ L. The split ratio is as follows: 10: 1; the temperature-raising program is as follows: the initial temperature is 40 ℃, the temperature is maintained for 5min, and the temperature is increased to 250 ℃ at the speed of 8 ℃/min.
Mass spectrum detection conditions: full scan mode monitoring, ion mass number scan range set to m/z 50-500, scan speed 1562[ N ═ 2], scan frequency 2.9/s.
(2) Quantitative analysis of health wine components
Establishing HPLC-QQQ-MS quantitative analysis method, and determining the content of 22 characteristic components in the medicinal materials of herba Epimedii, radix Puerariae, Cistanchis herba, radix astragali, Saviae Miltiorrhizae radix, fructus Lycii, Achyranthis radix, radix Angelicae sinensis, etc.
Chromatographic conditions are as follows: the column temperature is 25 ℃; mobile phase: water (containing a certain concentration of formic acid) (phase A) -acetonitrile (containing a certain concentration of formic acid) (phase B); flow rate: 0.3 mL/min; sample introduction volume: 5 μ L. The gradient elution conditions were as follows: 0-3.5min, 25-40% B; 3.5-4min, 40-75% B; 4-15min, 75% B.
Mass spectrum conditions: temperature of the drying gas: 350 ℃; flow rate of drying gas: 9L/min; carrier gas pressure: 30 psi; capillary voltage: 4000V (positive), 3500V (negative). Multiple Reaction Monitoring (MRM) mode detection in positive and negative ion mode.
And establishing a GC-MS quantitative analysis method to determine the contents of the borneol acetate and the eugenol in the health-care wine.
Chromatographic conditions are as follows: the instrument comprises the following steps: agilent 7890B GC-5977A MSD; a chromatographic column: agilent19091S-433UI HP-5ms Ultra Inert (30 m. times.250 μm,0.25 μm); 0-325 ℃; carrier gas: 99.999% high-purity He; flow rate: 1 mL/min; sample introduction volume: 1 μ L. The split ratio is as follows: 10: 1; the temperature-raising program is as follows: the initial temperature is 40 ℃, the temperature is maintained for 5min, and the temperature is increased to 250 ℃ at the speed of 8 ℃/min.
Mass spectrum conditions: ion source temperature: 320 ℃, quadrupole temperature: scanning is carried out under a single ion detection Scanning (SIM) mode at 150 ℃, and the highest abundance fragment ions of each detected substance are respectively used as a quantitative basis and are respectively an internal standard (methyl salicylate): m/z 120.1; bornyl acetate: m/z 95.2; eugenol: m/z is 164.1. Solvent retardation: for 3 minutes.
(3) Zebra fish model drug effect evaluation
The method is carried out in a six-well plate, a normal control group and a model group are arranged, 60-tail 4dpf AB-series zebra fish are treated in each experimental group, and a fatigue model is induced by sodium sulfite. The contents of lactic acid and ATP of zebra fish are detected after 3 days of treatment by using health care wine with different administration doses.
(4) Screening of anti-fatigue main effect component group
The importance of each component to the drug effect is comprehensively reflected by a specific mathematical expression by fitting a mathematical equation between the drug effect index and the content of each compound by adopting a partial least squares regression analysis method. The content data of 33 compounds and anti-inflammatory 2 pharmacodynamic indexes (lactic acid and ATP) measured in each batch of the health-care wine are input into Simca-P13.0 software for analysis, the content data of the compounds are independent variables X (X1, X2, X3, X4, X5, X6, X7, X8, X9 and X10), the lactic acid is dependent variable Y1, and the ATP is dependent variable Y2, and after PLS-DA analysis, a regression equation between the 2 pharmacodynamic indexes and the 33 compound contents of the health-care wine is obtained, and a regression coefficient graph is drawn. Mathematical statistics studies show that when conditions are met, regression coefficients obtained in the regression equation are used to predict the correlation between the study model and the efficacy index to a certain extent.
The method for establishing the quality control of the anti-fatigue components of the health-care wine by qualitatively and quantitatively analyzing the components of the health-care wine, evaluating the anti-fatigue drug effect of the zebra fish model and screening the anti-fatigue main active component group has the following advantages:
(1) the invention adopts HPLC-QTOF-MS/MS and GC-MS technologies to comprehensively analyze the components of the health-care wine, and discovers 33 volatile components and 89 compounds in total.
(2) The method adopts an HPLC-QQQ-MS quantitative analysis method and a GC-MS quantitative analysis method to carry out quantitative analysis on the monitorable components of the health-care wine, and realizes the quality control of the health-care wine by detecting 22 characteristic components and 2 volatile characteristic components in the health-care wine.
(3) The anti-fatigue effect of the health-care wine is evaluated through the zebra fish model, and the anti-fatigue main active ingredient group is screened through the least square method, so that the quality control of the anti-fatigue ingredients of the health-care wine is realized.
Drawings
FIG. 1 is a plot of PLS regression coefficients between lactic acid and 33 compound levels;
FIG. 2 is a plot of the PLS regression coefficients between ATP and 33 compound levels.
Detailed Description
The technical solution of the present invention is further explained by the following specific examples.
The health wine is prepared from Chinese medicinal materials with nourishing and treating functions and wine, belongs to one kind of compound wine, and has the functions of preventing and treating diseases and strengthening body. In the quality control of health wine, different detection means are adopted to ensure the stability of various component indexes of the wine. In the prior art, no quality control method for determining the anti-fatigue efficacy of the health care wine through animal experiments appears.
Example 1
The screening method of the anti-fatigue components of the health wine comprises the following steps: qualitative analysis of health wine components, quantitative analysis of health wine components, evaluation of zebra fish model drug effect, and screening of anti-fatigue main active ingredient groups.
Example 2
(1) Qualitative analysis of health wine ingredients in the present application
The HPLC-QTOF-MS/MS technology is adopted to carry out systematic identification research on chemical components contained in the health-care wine.
Chromatographic conditions are as follows: the column temperature is 25 ℃; mobile phase: water (containing formic acid with certain concentration) (A) -acetonitrile (containing formic acid with certain concentration) (B); flow rate: 0.8 mL/min; sample introduction volume: 2 μ L. The gradient elution conditions were: 0-2.5min, 10% B; 2.5-20min, 10-20% B; 20-35min, 20-35% B; 35-40min, 35-60% B; 40-45min, 60-90% B; 45-50min, 90-95% B; 50-60min, 95% B.
Mass spectrum conditions: flow rate of dry nitrogen: 10L/min; temperature of dry nitrogen: 350 ℃; atomization pressure: 35 psig; temperature of sheath gas: 350 ℃; flow rate of sheath gas: 11.0L/min; capillary voltage: 4000V; taper hole voltage: 65V; OCT 1RF Vpp: 750V; cleavage voltage: 135V; secondary collision energy: 15V, 25V, 35V and 50V. And (3) monitoring in a full scanning mode in a positive ion mode and a negative ion mode, wherein the scanning range of the mass number of the ions is set to be m/z 50-1700.
Through comparison with reference substances and literature data, 89 compounds are identified in total, wherein the 89 compounds comprise 33 flavonoids, 20 phenolic acids and organic acids, 8 triterpene saponins, 5 alcohol glycosides, 4 steroids, 3 saccharides, 3 alkaloids, 3 aldehydes, 2 iridoids, 2 phenylpropanoids, 2 amino acids, 2 amides, 1 coumarin and 1 nucleoside; carrying out medicinal material attribution on the identified chemical components, and finding that related components from epimedium and kudzu root are more, namely 25 and 18 compounds respectively; the characteristic components of the 22 traditional Chinese medicines are detected in the health wine.
The GC-MS technology is adopted to systematically identify volatile components from angelica, fructus amomi, rhizoma curculiginis, fructus alpiniae oxyphyllae, cinnamon and clove in the health care wine.
Chromatographic conditions are as follows: the instrument comprises the following steps: agilent 7890B GC-5977A MSD; a chromatographic column: agilent19091S-433UI HP-5ms Ultra Inert (30 m. times.250 μm,0.25 μm); 0-325 ℃; carrier gas: 99.999% high-purity He; flow rate: 1 mL/min; sample introduction volume: 1 μ L. The split ratio is as follows: 10: 1; the temperature-raising program is as follows: the initial temperature is 40 ℃, the temperature is maintained for 5min, and the temperature is increased to 250 ℃ at the speed of 8 ℃/min.
Mass spectrum detection conditions: full scan mode monitoring, ion mass number scan range set to m/z 50-500, scan speed 1562[ N ═ 2], scan frequency 2.9/s.
33 volatile components are identified from the health wine and the base wine, wherein 12 of the volatile components are selected from 6 medicinal materials such as angelica, fructus amomi, clove, fructus alpiniae oxyphyllae, cinnamon, rhizoma curculiginis and the like, and the rest is inherent components of the wine body; the borneol acetate in the fructus amomi and the eugenol in the clove can be used as characteristic volatile components of the health care wine.
(2) Quantitative analysis of health wine ingredients in the present application
Establishing HPLC-QQQ-MS quantitative analysis method, and determining the content of 22 characteristic components in the medicinal materials of herba Epimedii, radix Puerariae, Cistanchis herba, radix astragali, Saviae Miltiorrhizae radix, fructus Lycii, Achyranthis radix, radix Angelicae sinensis, etc.
Chromatographic conditions are as follows: the column temperature is 25 ℃; mobile phase: water (containing a certain concentration of formic acid) (phase A) -acetonitrile (containing a certain concentration of formic acid) (phase B); flow rate: 0.3 mL/min; sample introduction volume: 5 μ L. The gradient elution conditions were as follows: 0-3.5min, 25-40% B; 3.5-4min, 40-75% B; 4-15min, 75% B.
Mass spectrum conditions: temperature of the drying gas: 350 ℃; flow rate of drying gas: 9L/min; carrier gas pressure: 30 psi; capillary voltage: 4000V (positive), 3500V (negative). Multiple Reaction Monitoring (MRM) mode detection in positive and negative ion mode.
The highest puerarin content detected from the health wine is 10-20 mug/mL, and the contents of epimedin C, icariin, epimedin B, betaine and daidzin are 3-10 mug/mL, and the contents of other components are 0.05-1.00 mug/mL. The content difference of the measured components in the three batches of health care wine samples is not great, which shows that the product quality consistency is good.
And establishing a GC-MS quantitative analysis method to determine the contents of the borneol acetate and the eugenol in the health-care wine.
Chromatographic conditions are as follows: the instrument comprises the following steps: agilent 7890B GC-5977A MSD; a chromatographic column: agilent19091S-433UI HP-5ms Ultra Inert (30 m. times.250 μm,0.25 μm); 0-325 ℃; carrier gas: 99.999% high-purity He; flow rate: 1 mL/min; sample introduction volume: 1 μ L. The split ratio is as follows: 10: 1; the temperature-raising program is as follows: the initial temperature is 40 ℃, the temperature is maintained for 5min, and the temperature is increased to 250 ℃ at the speed of 8 ℃/min.
Mass spectrum conditions: ion source temperature: 320 ℃, quadrupole temperature: scanning is carried out under a single ion detection Scanning (SIM) mode at 150 ℃, and the highest abundance fragment ions of each detected substance are respectively used as a quantitative basis and are respectively an internal standard (methyl salicylate): m/z 120.1; bornyl acetate: m/z 95.2; eugenol: m/z is 164.1. Solvent retardation: for 3 minutes.
Detecting the contents of bornyl acetate and eugenol in the health care wine to be 5-20 mug/mL; the contents of the two components in three batches of health wine samples are not greatly different, which shows that the product quality consistency is good.
(3) Zebra fish model validation in the present application
The method is carried out in a six-well plate, a normal control group and a model group are arranged, 60-tail 4dpf AB-series zebra fish are treated in each experimental group, and a fatigue model is induced by sodium sulfite. The lactic acid and ATP contents of the zebra fish were measured after 3 days of treatment. The result shows that when the dilution multiple of the health-care wine is 800, the lactic acid content (p is less than 0.001) in the zebra fish body can be obviously reduced, and the ATP content (p is less than 0.001) in the zebra fish body is obviously increased, which indicates that the health-care wine has a certain improvement effect on the fatigue state of the zebra fish.
(4) Anti-fatigue major ingredient group screening in the present application
The importance of each component to the drug effect is comprehensively reflected by a specific mathematical expression by fitting a mathematical equation between the drug effect index and the content of each compound by adopting a partial least squares regression analysis method. The content data of 33 compounds and anti-inflammatory 2 pharmacodynamic indexes (lactic acid and ATP) measured in each batch of the health-care wine are input into Simca-P13.0 software for analysis, the content data of the compounds are independent variables X (X1, X2, X3, X4, X5, X6, X7, X8, X9 and X10), the lactic acid is dependent variable Y1, and the ATP is dependent variable Y2, and after PLS-DA analysis, a regression equation between the 2 pharmacodynamic indexes and the 33 compound contents of the health-care wine is obtained, and a regression coefficient graph is drawn. Mathematical statistics studies show that when conditions are met, regression coefficients obtained in the regression equation are used to predict the correlation between the study model and the efficacy index to a certain extent.
The invention discloses a method for screening anti-fatigue components of health-care wine, which comprises the following steps: (1) carrying out qualitative analysis on the components of the health wine; (2) carrying out quantitative analysis on characteristic components of the health-care wine; (3) evaluating the anti-fatigue efficacy by using a zebra fish model; (4) and screening the anti-fatigue main effect component group by using a least square method. Inputting the content data of 33 compounds selected by the health-care wine and the data of 2 anti-inflammatory drug effect indexes (lactic acid and ATP) into Simca-P13.0 software for analysis, obtaining a regression equation between the 2 drug effect indexes of the health-care wine and the 33 compound content, and obtaining a drug effect contribution value of the related coefficient judgment component. The method has the following advantages: (1) comprehensive qualitative analysis is carried out on the components of the health-care wine by adopting HPLC-QTOF-MS/MS and GC-MS technologies; (2) quantitative analysis is carried out on the monitorable components of the health-care wine by adopting an HPLC-QQQ-MS quantitative analysis method and a GC-MS quantitative analysis method; (3) the anti-fatigue effect of the health-care wine is evaluated through the zebra fish model, and the anti-fatigue main active ingredient group is screened through the least square method, so that the quality control of the anti-fatigue ingredients of the health-care wine is realized.
As shown in fig. 1, the regression equation between lactic acid and 33 chemical components content: y ═ 0.0014X + 0.0023X-0.0034X-0.0023X-0.0096X-0.0036X +0.0056X + 0.0004X-0.0045X-0.0037X +0.0002X +0.0126X +0.0056X +0.0088X +0.0035X +0.0176X +0.0074X + 0.0089X-0.0046X + 0.0056X-0.0002X-0.0075X-0.0130X-0.0184X-0.0070X-0.0055X-0.0041X +0.0002X +0.0004X +0.0031X +0.0116X + X.
As shown in fig. 2, the regression equation between ATP and 33 chemical components content: y ═ 0.0014X + 0.0023X-0.0034X-0.0023X-0.0096X-0.0036X +0.0056X + 0.0004X-0.0045X-0.0037X +0.0002X +0.0126X +0.0056X +0.0088X +0.0035X +0.0176X +0.0074X + 0.0089X-0.0046X + 0.0056X-0.0002X-0.0075X-0.0130X-0.0184X-0.0070X-0.0055X-0.0041X +0.0002X +0.0004X +0.0031X +0.0116X + X.
The compounds A23 (daidzin), A24 (puerarin), A25 (formononetin), A26 (calycosin glucoside), A27 (icariside II), A10 (p-coumaric acid), A4 (baohuoside II) and A21 (daidzein) all have drug effect contributions to 2 drug effect indexes, so that the 8 compounds are considered as main active ingredient groups for playing the anti-fatigue drug effect.
The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (5)
1. A method for screening anti-fatigue components of health wine is characterized by comprising the following steps: qualitative analysis of health wine components, quantitative analysis of health wine components, evaluation of zebra fish model drug effect, and screening of anti-fatigue main active ingredient groups.
2. The method for screening the anti-fatigue components of the health-care wine according to claim 1, wherein the qualitative analysis of the components of the health-care wine comprises the following steps:
identifying chemical components contained in the health-care wine by adopting an HPLC-QTOF-MS/MS technology:
chromatographic conditions are as follows: the column temperature is 25 ℃; mobile phase: water (containing formic acid with certain concentration) (A) -acetonitrile (containing formic acid with certain concentration) (B); flow rate: 0.8 mL/min; sample introduction volume: 2 mu L of the solution; the gradient elution conditions were: 0-2.5min, 10% B; 2.5-20min, 10-20% B; 20-35min, 20-35% B; 35-40min, 35-60% B; 40-45min, 60-90% B; 45-50min, 90-95% B; 50-60min, 95% B;
mass spectrum conditions: flow rate of dry nitrogen: 10L/min; temperature of dry nitrogen: 350 ℃; atomization pressure: 35 psig; temperature of sheath gas: 350 ℃; flow rate of sheath gas: 11.0L/min; capillary voltage: 4000V; taper hole voltage: 65V; OCT 1RF Vpp: 750V; cleavage voltage: 135V; secondary collision energy: 15V, 25V, 35V, 50V; monitoring in a full scanning mode in a positive ion mode and a negative ion mode, wherein the scanning range of the ion mass number is set to be m/z 50-1700;
identifying volatile components from angelica, fructus amomi, rhizoma curculiginis, fructus alpiniae oxyphyllae, cinnamon and clove in the health care wine by adopting GC-MS technology:
chromatographic conditions are as follows: the instrument comprises the following steps: agilent 7890B GC-5977A MSD; a chromatographic column: agilent19091S-433UI HP-5ms Ultra Inert (30 m. times.250 μm,0.25 μm); 0-325 ℃; carrier gas: 99.999% high-purity He; flow rate: 1 mL/min; sample introduction volume: 1 mu L of the solution; the split ratio is as follows: 10: 1; the temperature-raising program is as follows: the initial temperature is 40 ℃, the temperature is maintained for 5min, and the temperature is increased to 250 ℃ at the speed of 8 ℃/min.
Mass spectrum detection conditions: full scan mode monitoring, ion mass number scan range set to m/z 50-500, scan speed 1562[ N ═ 2], scan frequency 2.9/s.
3. The method for screening the anti-fatigue components of the health-care wine according to claim 1, wherein the quantitative analysis of the components of the health-care wine comprises:
establishing HPLC-QQQ-MS quantitative analysis method, and determining content of characteristic components in the health wine from herba Epimedii, radix Puerariae, Cistanchis herba, radix astragali, Saviae Miltiorrhizae radix, fructus Lycii, Achyranthis radix, and radix Angelicae sinensis;
chromatographic conditions are as follows: the column temperature is 25 ℃; mobile phase: water (containing a certain concentration of formic acid) (phase A) -acetonitrile (containing a certain concentration of formic acid) (phase B); flow rate: 0.3 mL/min; sample introduction volume: 5 mu L of the solution; the gradient elution conditions were as follows: 0-3.5min, 25-40% B; 3.5-4min, 40-75% B; 4-15min, 75% B;
mass spectrum conditions: temperature of the drying gas: 350 ℃; flow rate of drying gas: 9L/min; carrier gas pressure: 30 psi; capillary voltage: 4000V (positive), 3500V (negative); detecting in a multiple reaction monitoring mode in a positive and negative ion mode;
establishing a GC-MS quantitative analysis method, and determining the contents of borneol acetate and eugenol in the health-care wine:
chromatographic conditions are as follows: the instrument comprises the following steps: agilent 7890B GC-5977A MSD; a chromatographic column: agilent19091S-433UI HP-5ms Ultra Inert (30 m. times.250 μm,0.25 μm); 0-325 ℃; carrier gas: 99.999% high-purity He; flow rate: 1 mL/min; sample introduction volume: 1 mu L of the solution; the split ratio is as follows: 10: 1; the temperature-raising program is as follows: the initial temperature is 40 ℃, the temperature is maintained for 5min, and the temperature is increased to 250 ℃ at the speed of 8 ℃/min;
mass spectrum conditions: ion source temperature: 320 ℃, quadrupole temperature: scanning is carried out under a single ion detection Scanning (SIM) mode at 150 ℃, and the highest abundance fragment ions of each detected substance are respectively used as a quantitative basis and are respectively an internal standard (methyl salicylate): m/z 120.1; bornyl acetate: m/z 95.2; eugenol: m/z is 164.1; solvent retardation: for 3 minutes.
4. The method for screening the anti-fatigue components of the health-care wine as claimed in claim 1, wherein the evaluation of the drug effect of the zebra fish model comprises the following steps:
setting a normal control group and a model group in a six-hole plate, treating 60-tail 4dpf AB-series zebra fish in each experimental group, and inducing a fatigue model by using sodium sulfite; the contents of lactic acid and ATP of zebra fish are detected after 3 days of treatment by using health care wine with different administration doses.
5. The method for screening the anti-fatigue components for the health-care wine according to claim 1, wherein the screening of the anti-fatigue main active component group comprises the following steps:
the importance of each component to the drug effect is comprehensively reflected by a specific mathematical expression by fitting a mathematical equation between the drug effect index and the content of each compound by adopting a partial least squares regression analysis method; inputting the content data of the compounds and anti-inflammatory 2 pharmacodynamic indexes (lactic acid and ATP) data measured in each batch of the health-care wine into Simca-P13.0 software for analysis, wherein the content data of the compounds are independent variables X (X1, X2, X3, X4, X5, X6, X7, X8, X9 and X10), the lactic acid is dependent variable Y1, the ATP is dependent variable Y2, and after PLS-DA analysis, regression equations between the 2 pharmacodynamic indexes and the compound content of the health-care wine are obtained, and a regression coefficient graph is drawn; mathematical statistics studies show that when conditions are met, regression coefficients obtained in the regression equation are used to predict the correlation between the model and the efficacy index.
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