CN112098560A - Method for quantitatively detecting total flavonoids at different parts of walnut by using coefficient conversion method - Google Patents
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Abstract
The invention relates to a method for quantitatively detecting total flavonoids at different parts of walnuts by a coefficient conversion method. The method mainly comprises the following steps: 1) preparing a reference substance solution; 2) preparing a test solution; 3) measuring the content of hydrolyzed main aglycone by UPLC-UV; 4) the total flavonoids were calculated by coefficient conversion. The detection method established by the invention has the advantages of simplicity, convenience, rapidness, high sensitivity, strong specificity and good repeatability, and is successfully applied to the content determination of the total flavonoids at different parts of the walnut. The invention not only provides a theoretical basis for formulating the walnut quality standard, provides key information for researchers and producers, but also provides a research foundation for better developing and utilizing walnut resources.
Description
Technical Field
The invention relates to a method for quantitatively detecting total flavonoids at different parts of walnuts by a coefficient conversion method.
Background
Juglans regia L is a kind of Juglans deciduous tree of Juglans of Juglandaceae, and can be used as medicine for walnut kernel, diaphragma juglandis, green tangerine orange peel, leaf, flower, shell, etc. The Chinese herbal medicine compilation records that walnut leaves, named as walnut leaves, are leaves of Juglans regia L.of Juglandaceae, are used for elephantiasis, leukorrhagia and mange, and are fresh and collected at any time.
Researches show that the walnut leaves contain high-content flavonoid substances which are good sources of the flavonoid compounds, the flavonoid compounds are main chemical components of the pericarpium juglandis, the flavonoid compounds in the diaphragma juglandis are one of the main components, and the walnut shells and the walnut kernels contain the flavonoid compounds. The flavone has stronger antioxidation, is closely related to the regulation of immunologic function and the enhancement of anticancer activity, and is necessary for the content determination of the total flavone of different parts of the walnut.
At present, the content determination of total flavonoids mostly adopts an aluminum nitrate-sodium nitrite colorimetric method, but impurities possibly have ultraviolet absorption at a set wavelength in the determination process of the method, so that the final result is influenced; in addition, the traditional Chinese medicine components are complex, the flavonoid glycoside components are difficult to obtain, and the total flavonoid can not be directly quantified by obtaining a reference substance of all flavonoid components.
Disclosure of Invention
Aiming at the defects of the prior art, the application provides a simple and easily-obtained method for quantitatively detecting the total flavonoids at different parts of the walnut by a coefficient conversion method.
A method for quantitatively detecting total flavonoids at different parts of walnuts by a coefficient conversion method comprises the following steps:
(1) preparing a reference substance solution;
(2) preparing a test solution;
(3) measuring the content of hydrolyzed main aglycone by UPLC-UV;
(4) the total flavonoids were calculated by coefficient conversion.
Further, the main aglycone in the step (3) comprises: quercetin and kaempferol.
Further, the preparation of the reference solution in step (1) comprises: precisely weighing a proper amount of quercetin and kaempferol standard substances, respectively dissolving with methanol, and preparing into independent standard stock solutions; respectively and precisely measuring appropriate amount of standard stock solution, and diluting with methanol to prepare mixed working solution; serially diluting the mixed working solution to obtain mixed standard series solutions of quercetin and kaempferol with the concentrations of 1, 2, 5, 10, 50, 100 and 250 mug/mL.
Further, the preparation of the test solution in the step (2) comprises: weighing 0.5g of a walnut sample to be detected, placing the walnut sample into a test tube with a plug, precisely weighing 10.00mL of 50% methanol, carrying out ultrasonic treatment in an ultrasonic cleaning instrument for 15min at room temperature, cooling, centrifuging for 10min by adopting a high-speed refrigerated centrifuge, taking 2.00mL of supernatant, carrying out rotary evaporation at 55 ℃, and cooling to room temperature; re-dissolving 2.00mL of pure methanol, adding 0.40mL of 37% hydrochloric acid, condensing, refluxing and hydrolyzing for 1h at 55 ℃, and cooling; adding 1.60mL of pure methanol, mixing, filtering with 0.22 μm filter membrane, and collecting the filtrate to obtain the sample solution.
Further, the content of the hydrolyzed main aglycone is determined by UPLC-UV in the step (4), wherein the liquid chromatography condition is as follows
Has the advantages that:
the sample pretreatment method provided by the invention is simple to operate. The preparation of the test solution only needs 50% methanol as an extraction solvent, and the ratio of the material to the liquid is 1: and (3) performing ultrasonic treatment at room temperature for 15min, centrifuging, taking a certain volume, performing rotary evaporation, adding methanol and hydrochloric acid for redissolving, and performing acid hydrolysis analysis. The extraction efficiency of the sample pretreatment process is high, and the interference of sample analysis is less;
the invention adopts the technology of combining the ultrasonic auxiliary extraction method and the acid hydrolysis method for the first time, and the ultrasonic auxiliary extraction method has high extraction efficiency, short extraction time and simple and easy operation; the acid hydrolysis method has less interference and simple operation. The method is adopted to extract the total flavonoids in different parts of the walnut, and the methodological verification is carried out;
the invention adopts the acid hydrolysis method to extract the total flavone in different parts of the walnut for the first time, and adopts the coefficient conversion method to carry out the content measurement of the total flavone in different parts of the walnut in different producing areas for the first time (because the flavone aglycones generated after the acid hydrolysis of different parts are possibly inconsistent, the invention firstly takes the walnut leaves as the main part to carry out the acid hydrolysis, carries out methodology verification after quantifying the aglycones generated by the acid hydrolysis, and then applies the method to quantify the total flavone in different parts of the walnut, the invention only aims at the part where the flavone aglycones generated after the acid hydrolysis are quercetin or kaempferol, and because the content of the flavone aglycone glucoside in different parts is different, the adopted conversion coefficient is also different when carrying out the content calculation);
according to the invention, the flavonoid glycoside in different parts of the walnut is hydrolyzed into flavonoid aglycone by adopting an acid hydrolysis method, so that the reasonable utilization of the walnut can be used as a theoretical and experimental basis, a reference basis is provided for establishing content measurement standards of different parts of the walnut, and the development and utilization of the walnut are facilitated;
the UPLC adopted by the invention has the advantages of simplicity, convenience, rapidness, high sensitivity, strong specificity and good repeatability, and can complete the detection of acid hydrolysis products of different parts of walnuts within 20 min. On the basis of establishing a standard curve, the content of main flavonoid aglycone generated by acid hydrolysis is measured to obtain rich information content.
Detailed Description
The equipment and raw materials used in the application are as follows:
methanol and acetonitrile are both chromatographically pure and purchased from merck corporation;
acetic acid was chromatographically pure, purchased from semer fly;
the experimental water is Wahaha water, and the hydrochloric acid is analytically pure.
Quercetin (CAS: 117-39-5), purchased from China pharmaceutical biologies institute;
kaempferol (CAS: 520-18-3), available from Shanghai Allan Biotechnology Ltd; all standards were 98% pure.
One-ten-thousandth balance (model: XPE 205): mettler corporation, Switzerland;
one-ten-thousandth balance (model: XPE 204): mettler corporation, Switzerland;
ultrasonic cleaner (model: SK250 HP): shanghai Kedao ultrasonic instruments, Inc.;
high speed refrigerated centrifuge (model: Sorvall ST 8R): sammer Feishel technologies, Inc.;
heat collection type constant temperature magnetic stirrer (model: DF-101S): consolidate City Prohua instruments, Inc.;
rotary evaporator (model: HB 10S 096): germany Aika company;
ultra high performance liquid chromatography (model: UPLC I-CLASS): vortish, USA.
Examples
The application relates to a method for quantitatively detecting total flavonoids at different parts of walnuts by a coefficient conversion method, which comprises the following steps:
(1) preparing a reference substance solution;
(2) preparing a test solution;
(3) measuring the content of hydrolyzed main aglycone by UPLC-UV;
(4) the total flavonoids were calculated by coefficient conversion.
As an embodiment of the present invention, the preparation of a control solution comprises: precisely weighing appropriate amount of quercetin and kaempferol standard substance, dissolving with methanol respectively, and making into independent standard stock solution. And precisely measuring appropriate amounts of the standard stock solutions respectively, and diluting with methanol to prepare a mixed working solution. Serially diluting the mixed working solution to obtain mixed standard series solutions of quercetin and kaempferol with concentrations of 1, 2, 5, 10, 50, 100 and 250 μ g/mL. All solutions were stored in a 4 ℃ freezer until analysis.
As an embodiment of the present invention, the preparation of a test solution comprises: weighing 0.5g of a walnut sample to be measured, precisely weighing, placing in a test tube with a plug, precisely weighing 10.00mL of 50% methanol, ultrasonically treating in an ultrasonic cleaner for 15min at room temperature, cooling, centrifuging for 10min by adopting a high-speed refrigerated centrifuge, taking 2.00mL of supernatant, rotatably steaming at 55 ℃, and cooling to room temperature; re-dissolving 2.00mL of pure methanol, adding 0.40mL of 37% hydrochloric acid, condensing, refluxing and hydrolyzing for 1h at 55 ℃, and cooling; adding 1.60mL of pure methanol, mixing, filtering with 0.22 μm filter membrane, and collecting the filtrate to obtain the sample solution. The method adopts an ultrasonic-assisted method to extract different parts of the walnut, adopts an acid hydrolysis method to hydrolyze flavonoid glycoside, and adopts a technology combining an ultrasonic-assisted extraction method and an acid hydrolysis method. The invention adopts the technology of combining the ultrasonic auxiliary extraction method and the acid hydrolysis method for the first time, and the ultrasonic auxiliary extraction method has high extraction efficiency, short extraction time and simple and easy operation; the acid hydrolysis method can hydrolyze the flavonoid glycoside in the medicinal material into flavonoid aglycone, and the content of the main flavonoid aglycone generated by acid hydrolysis can be more accurately reflected.
The sample pretreatment method is important, and the sample pretreatment is a key link of quantitative analysis of active ingredients. The pretreatment of the invention is examined in two parts, and the first part is the extraction of total flavonoids in different parts of the walnut. The extraction solvents of 25%, 50%, 75% and 100% methanol were considered, respectively, with a feed-to-liquid ratio of 1: 10. 1: 20. 1: 50 and 1: 100 and peak areas with ultrasonic time of 0, 15, 30, 45min, and the extraction conditions were found to be 50% methanol, 1 feed-to-liquid ratio: the extraction efficiency is highest at 20min and 15 min. The second part is a review of the hydrolysis conditions. Methanol-hydrochloric acid ratio (V/V) at hydrolysis was examined to be 6: 1. 5: 1. 4: 1, the sum of peak areas of quercetin and kaempferol at hydrolysis temperatures of 45 ℃, 55 ℃ and 70 ℃ and hydrolysis times of 45, 60 and 120min, and the hydrolysis conditions were found to be methanol-hydrochloric acid 5: 1. the hydrolysis temperature is 55 ℃ and the hydrolysis time is 60 min.
Determination of content
Quality of main flavonoid aglycone after hydrolysis
In the formula (1), the reaction mixture is,
Xi-mass (mg) of quercetin or kaempferol in the sample
CiFrom a standard curveThe concentration of Quercetin or Kaempferol in the sample (μ g/mL)
V0Volume of extraction solvent (mL)
V1Volume of supernatant (mL) obtained by rotary evaporation
V2Total volume of sample after hydrolysis (mL)
In the formula (2), the reaction mixture is,
y-content of Total Flavonoids in different parts of walnut (%)
Xi-mass (mg) of quercetin or kaempferol after hydrolysis of different parts of the walnut
TiConversion factor between aglycone molecular weight and glycoside molecular weight (see Table 2)
m-sample weighing (g)
TABLE 1 conversion coefficient of Quercetin to Kaempferol
The content of quercetin hexaglucoside in the walnut leaves is obviously higher than that of other glycosides of quercetin, so that the quercetin hexaglucoside is adopted; flavone glycosides (namely kaempferol pentoside and kaempferol hexoside) taking kaempferol as aglycone in walnut leaves and flavone glycosides (namely quercetin pentoside and quercetin hexoside) taking quercetin as aglycone in Qinglongyi and Xinjiang nut, because the content of the corresponding flavone glycosides can not be directly determined, pentoside is selected, namely the result obtained by calculation is the lowest flavone content; and (4) combining the above steps, and calculating the result to be the lowest total flavone content in the sample.
The content determination methodology was verified as follows:
linear range, detection limit, quantitation limit:
taking mixed standard series solution, treating according to the above sample treatment method, injecting sample, and measuring, wherein each component isThe peak area of (A) is subjected to linear regression on the concentration of the (B) with a weight factor of 1/X2And obtaining a corresponding regression equation. The regression equation of quercetin is Y-55400X-110000 (r-0.9997), and the result shows that the linear relation of quercetin is good in the linear range of 1-250 mug/mL; the regression equation for kaempferol is Y72000X-18900 (r 0.9999), and the results show that kaempferol has good linear relationship in the linear range of 1-250 μ g/mL, and the results are shown in Table 2.
Respectively measuring appropriate amount of mixed standard stock solution, diluting with methanol, performing sample injection measurement according to the above detection conditions, and recording chromatogram. Taking the sample injection concentration when the peak response value is 10 times of the baseline noise (10 times of the signal-to-noise ratio) as a quantitative Limit (LOQ); the injection concentration at which the peak response was 3 times the baseline noise (3 times the signal-to-noise ratio) was taken as the limit of detection (LOD), and the results are shown in Table 2.
TABLE 2 regression equation, correlation coefficient, linear range, detection limit and quantitation limit for control quercetin and kaempferol
Precision of the instrument:
taking a walnut leaf sample solution, carrying out continuous sample injection for 6 times according to the chromatographic conditions, respectively recording peak areas of quercetin and kaempferol, calculating that the RSD values are 0.43 percent and 0.35 percent respectively, and indicating that the precision of the instrument is good.
And (3) standard addition recovery rate:
about 0.5g of the same batch of samples with known content is precisely weighed, 9 parts of the samples are respectively added with three reference substance amounts of low, medium and high 3 levels (80%, 100% and 120%), the samples are processed according to the sample processing method and are measured in parallel, and the result shows that the accuracy is good. The results are shown in Table 3.
TABLE 3 results of recovery with addition of standard
Repeatability:
about 0.5g of the same batch of samples are precisely weighed and paralleled in six parts, and the test solution is prepared and paralleled for measurement according to the method. The RSD values of quercetin and kaempferol are calculated to be 0.44% and 0.50%, respectively, which indicates that the method has good repeatability. The results are shown in Table 4.
Stability:
taking the same batch of samples, preparing into test solution by the method, and measuring the content of each component in 0, 2, 4, 8, 12 and 24h respectively. The RSD values of quercetin and kaempferol were calculated to be 3.76% and 2.42%, respectively, indicating good stability, and the results are shown in table 4.
TABLE 4 repeatability and stability results
| Composition (I) | Repeated (n ═ 6) RSD/%) | Stability (n-6, 24h) RSD/%) |
| Quercetin | 0.44 | 3.76 |
| Kaempferol | 0.50 | 2.42 |
As an example of the present invention, the content of the hydrolyzed aglycon was measured by UPLC-UV, wherein the liquid chromatography conditions are shown in Table 5 below.
TABLE 5 liquid chromatography conditions
The selection of the chromatographic column is important, and the Waters ACQUITY UPLC HSS T3 chromatographic column belongs to a hydrophilic ultrahigh pressure chromatographic column, and can realize better separation in a short time, and has better peak shape and stronger compound response. The flavonoid compounds have high polarity, and have poor separation effect and poor peak shape on a common C18 chromatographic column, and have better separation effect on a hydrophilic chromatographic column. In the experiment, Waters ACQUITY UPLC BEH C18, HSS PFP chromatographic columns and the like are tried, and the conditions that the peak shape is poor and the peak tailing occurs are found, so that the measurement is influenced. Therefore, a Waters ACQUITY UPLC HSS T3 column was finally selected for the assay.
In the present invention, the optimization of the mobile phase and gradient conditions is important, and other components in the sample may interfere with the detection of the target component during quantitative analysis, so that the elution conditions need to be optimized to ensure that the quantitative analysis of quercetin and kaempferol is performed in as short a time as possible. Therefore, the invention is optimized to determine the mobile phase to be 0.2% acetic acid water solution (phase A) -acetonitrile (phase B), and the gradient elution is carried out for 15min of running time.
The invention adopts a hydrolysis method and proper hydrolysis conditions to hydrolyze the flavonoid glycoside into corresponding flavonoid aglycone, measures the hydrolyzed flavonoid aglycone, and realizes the content measurement of the total flavonoid at different parts of the walnut by utilizing the conversion coefficient of the flavonoid aglycone and the flavonoid glycoside. Specifically, the method quantitatively analyzes the main flavonoid aglycone of the hydrolyzed walnut leaf medicinal material: quercetin and kaempferol. The medicinal materials contain two components of flavonoid glycoside and flavonoid aglycone, firstly, the flavonoid glycoside is converted into the flavonoid aglycone by adopting acid hydrolysis, then the content of each aglycone is determined, and finally, the content of the total flavonoid is calculated by coefficient conversion according to the molecular weight relationship between the glycoside and the corresponding aglycone. The content of the total flavonoids in different parts of the walnut is measured by adopting a coefficient conversion method, so that the measuring method for measuring the total flavonoids can be simplified, and the total flavonoids can be quantified only by obtaining a reference substance of main flavonoid aglycones after hydrolysis. The experimental method is simple and feasible, and simultaneously overcomes the defect that the flavonoid glycoside reference substance is not easy to obtain.
Application example
Content determination of total flavonoids in different parts of walnut
The analysis is carried out on 15 batches of samples of different parts of walnuts from different sources: the sample solution was prepared according to the sample treatment method of example 1, and the content measurement was performed according to the verified method, and the measurement results are shown in table 6.
TABLE 6 content measurement results of quercetin and kaempferol after hydrolysis of different parts of walnut
Note: "< LQD" indicates that the concentration is below the detection limit; "-" indicates that the component was not detected.
The quercetin and kaempferol detected in the above table are the components with the highest flavonoid aglycone content obtained after the hydrolysis of walnut leaves, while the main flavonoid aglycone obtained after the hydrolysis of exocarpium Juglandis Immaturum, diaphragma juglandis and walnut shells is quercetin, and the walnut kernel is reported in literature to also contain flavonoid components, and the content is not detected to be possibly too low. In walnut leaves in different producing areas, the content of the hydrolyzed quercetin is higher than that of the kaempferol. The content of total flavone in exocarpium Juglandis Immaturus and ramulus et folium Baphicacanthis Cusiae is obviously lower than that of folium Juglandis. Based on the results, the sequence of the flavone contents of different parts of the walnut is as follows: walnut leaves > diaphragma juglandis ≈ green dragon skin > walnut shell > walnut kernel. The flavonoid substance is one of anti-inflammatory and anti-tumor active ingredients of known medicines, and as the flavonoid glycoside reference substance is not easy to obtain, the flavonoid glycoside in different parts of the walnut is subjected to acid hydrolysis to obtain flavonoid aglycone, then the content of each aglycone is determined, and finally the content of the total flavonoid is calculated through coefficient conversion according to the molecular weight relationship between the glycoside and the corresponding aglycone. The content of the total flavonoids in different parts of the walnut is measured by adopting a coefficient conversion method, so that the measuring method for measuring the total flavonoids can be simplified, and the total flavonoids can be quantified only by obtaining a reference substance of main flavonoid aglycones after hydrolysis.
The invention firstly adopts a coefficient conversion method to carry out content measurement on the total flavone in different producing areas of different parts of the walnut (the flavonoid glycoside is hydrolyzed by acid to generate flavonoid aglycone, the molecular weights of the flavonoid glycoside and the aglycone are different, the conversion coefficient of the molecular weight of the main aglycone and the molecular weight of the glucoside in the different parts of the walnut is calculated, and the method is applied to the quantification of the total flavone).
The present invention may be embodied in other specific forms without departing from its spirit or essential characteristics. The above-described embodiments of the invention are therefore to be considered in all respects as illustrative and not restrictive, the scope of the invention being indicated by the appended claims, and not by the foregoing description, and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein.
Claims (5)
1. A method for quantitatively detecting total flavonoids at different parts of walnuts by a coefficient conversion method is characterized by comprising the following steps:
(1) preparing a reference substance solution;
(2) preparing a test solution;
(3) measuring the content of hydrolyzed main aglycone by UPLC-UV;
(4) the total flavonoids were calculated by coefficient conversion.
2. The method of claim 1, wherein the aglycon of step (3) comprises: quercetin and kaempferol.
3. The method of claim 1, wherein the preparation of the control solution in step (1) comprises: precisely weighing a proper amount of quercetin and kaempferol standard substances, respectively dissolving with methanol, and preparing into independent standard stock solutions; respectively and precisely measuring appropriate amount of standard stock solution, and diluting with methanol to prepare mixed working solution; serially diluting the mixed working solution to obtain mixed standard series solutions of quercetin and kaempferol with the concentrations of 1, 2, 5, 10, 50, 100 and 250 mug/mL.
4. The method of claim 1, wherein the preparing of the test solution in step (2) comprises: weighing 0.5g of a walnut sample to be detected, placing the walnut sample into a test tube with a plug, precisely measuring 10mL of 50% methanol, carrying out ultrasonic treatment in an ultrasonic cleaning instrument for 15min at room temperature, cooling, centrifuging for 10min by adopting a high-speed refrigerated centrifuge, taking 2.00mL of supernatant, carrying out rotary evaporation at 55 ℃, and cooling to room temperature; re-dissolving 2.0mL of pure methanol, adding 0.40mL of 37% hydrochloric acid, condensing, refluxing and hydrolyzing at 55 ℃ for 1h, and cooling; adding 1.60mL of pure methanol, mixing, filtering with 0.22 μm filter membrane, and collecting the filtrate to obtain the sample solution.
5. The method of claim 1, wherein the content of the hydrolyzed aglycon is determined by UPLC-UV in the step (4), and the liquid chromatography conditions are as follows
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107607654A (en) * | 2017-11-09 | 2018-01-19 | 中国检验检疫科学研究院 | The analysis method of flavonoids in a kind of walnut flower |
| WO2018130091A1 (en) * | 2017-01-13 | 2018-07-19 | 江南大学 | Method for extracting polyphenol substances from semen juglandis with inner seed coats by aid of combined enzyme and ultrasonic treatment |
-
2020
- 2020-09-22 CN CN202010999946.9A patent/CN112098560A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2018130091A1 (en) * | 2017-01-13 | 2018-07-19 | 江南大学 | Method for extracting polyphenol substances from semen juglandis with inner seed coats by aid of combined enzyme and ultrasonic treatment |
| CN107607654A (en) * | 2017-11-09 | 2018-01-19 | 中国检验检疫科学研究院 | The analysis method of flavonoids in a kind of walnut flower |
Non-Patent Citations (7)
| Title |
|---|
| GRAY, DE 等: "Quantitative analysis of flavonol glycosides in Ginkgo biloba: a comparison of two analytical methods", 《PHYTOCHEMICAL ANALYSIS》 * |
| 丁震 等: "《中药学（士、师、中级）应试指导及历年考点串讲》", 31 January 2019, 北京航空航天大学出版社 * |
| 任冰如 等: "HPLC测定红凤菜总黄酮含量", 《中国野生植物资源》 * |
| 任珊珊 等: "中药中总黄酮的含量测定方法研究进展", 《北方药学》 * |
| 孙奕 等: "近缘植物仙鹤草、委陵菜和翻白草中6种黄酮类化合物含量的比较", 《中药材》 * |
| 文红梅 等: "HPLC法测定蒲黄中总黄酮的含量", 《现代中药研究与实践》 * |
| 赵磊 等: "核桃花中槲皮素与山萘酚含量的高效液相色谱-二极管阵列检测器法测定", 《时珍国医国药》 * |
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Application publication date: 20201218 |