Flavonoids in phellodendri cortex and extraction method thereof

Abstract

This invention belongs to the field of agricultural product processing technology, and specifically relates to a flavonoid substance from wampee and its extraction method. A method for extracting flavonoid substances from wampee includes the following steps: (1) pulping wampee, adding a compound enzyme, subjecting it to a first ultra-high pressure treatment and enzymatic hydrolysis to obtain enzymatically hydrolyzed wampee pulp; (2) adding the enzymatically hydrolyzed wampee pulp obtained in step (1) to an extraction solvent, then adding an alkaline solution, subjecting it to a second ultra-high pressure treatment to obtain a wampee pulp treated with a second ultra-high pressure; (3) adding a solvent to the wampee pulp obtained in step (2), then adding an acid solution, performing solid-liquid separation to obtain flavonoid substances; the compound enzyme includes at least two of pectinase, cellulase, and hemicellulase. This invention not only promotes the conversion of insoluble bound flavonoid substances into free flavonoid substances, significantly improving the extraction yield of flavonoid substances and enriching the composition of flavonoid compounds, but also improves extraction efficiency and has good application value.

Description

Technical Field

[0001] This invention belongs to the field of agricultural product processing technology, and specifically relates to a flavonoid substance from wampee and its extraction method. Background Technology

[0002] Wampee is a dual-purpose resource for both medicinal and culinary uses, and is one of the ten finest fruits of Lingnan (southern China). It is mainly distributed in Guangdong, Guangxi, Hainan, Fujian, and Taiwan. Currently, wampee is primarily consumed fresh, while processed products include dried fruit and candied fruit. Traditional Chinese medicine records that wampee has digestive and cough-suppressing effects. Existing research indicates that wampee is rich in functional components such as flavonoids. Flavonoids are secondary metabolites widely found in plants, possessing antioxidant and anti-inflammatory properties, and are widely used in food and daily chemical products.

[0003] Most flavonoids have low solubility in water but high solubility in organic solvents such as methanol and ethanol, as well as under alkaline conditions. Currently, the main methods for flavonoid extraction include hot water extraction, organic solvent extraction, and ultrasound-assisted extraction. Industrial production of flavonoids from traditional Chinese medicine widely uses organic solvent extraction, which is time-consuming. Unlike traditional Chinese medicine, *Clausena lansium* (a type of medicinal herb) is rich in polysaccharides and cellulose. During hot water extraction, pectin and cellulose easily form gels and encapsulate flavonoids. In organic solvents, pectin and cellulose easily co-precipitate with flavonoids, making it difficult for flavonoids to dissolve and resulting in a low extraction rate. Ultrasonic-assisted extraction can improve the efficiency of flavonoid extraction, slightly increasing the extraction rate, but the overall flavonoid extraction rate remains low. Furthermore, some flavonoids in *Clausena lansium* easily cross-link with polysaccharides, lignin, and other components, forming insoluble bound flavonoids that are difficult to extract, further contributing to the low yield of flavonoids.

[0004] Therefore, there is an urgent need to provide a method for extracting flavonoids from yellow peel. This method can not only promote the conversion of insoluble bound flavonoids into free flavonoids, significantly improve the extraction yield of flavonoids, enrich the composition of flavonoid compounds, but also improve extraction efficiency and has good application value. Summary of the Invention

[0005] This invention aims to solve one or more technical problems existing in the prior art, and at least provide a beneficial option or create conditions. This invention provides a method for extracting flavonoids from *Clausena lansium*, which not only promotes the conversion of insoluble bound flavonoids into free flavonoids, significantly improving the extraction yield and enriching the composition of flavonoid compounds, but also improves extraction efficiency and has good application value.

[0006] The inventive concept of this invention: This invention employs a combination of enzymatic, ultra-high pressure (UHAP) treatment, and alkaline methods to extract flavonoids from wampee. The complex enzyme hydrolyzes pectin and cellulose in wampee, reducing their impact on flavonoid extraction. The first UHAP treatment not only activates the complex enzyme but also ensures sufficient contact between the enzyme and pectin and cellulose in the wampee, improving enzymatic hydrolysis efficiency. Simultaneously, the first UHAP treatment alters the structure of pectin, making it easier to decompose. The second UHAP treatment ensures sufficient contact between the extraction solvent and the wampee pulp, increasing the flavonoid extraction yield and shortening the extraction time. Furthermore, by adding alkaline solution to adjust the pH of the system to an alkaline state, insoluble bound flavonoids can be hydrolyzed and dissolved. The combined effect of the alkali and the second UHAP treatment increases the solubility, thereby improving the flavonoid extraction yield.

[0007] Therefore, a first aspect of the present invention provides a method for extracting flavonoids from yellow peel.

[0008] Specifically, a method for extracting flavonoids from yellow peel includes the following steps:

[0009] (1) Pulp the yellow peel, add compound enzyme, and then perform a first ultra-high pressure treatment and enzymatic hydrolysis to obtain enzymatic hydrolyzed yellow peel pulp;

[0010] (2) Add the enzymatically hydrolyzed yellow peel pulp obtained in step (1) to the extraction solvent, then add alkaline solution, and then perform a second ultra-high pressure treatment to obtain yellow peel pulp treated with two ultra-high pressures.

[0011] (3) Add the solvent to the yellow peel pulp obtained in step (2), then add acid, and separate the solid and liquid to obtain flavonoids;

[0012] In step (1), the complex enzyme includes at least two of pectinase, cellulase, and hemicellulase.

[0013] Preferably, in step (1), the wampee includes at least one of frozen wampee fruit and fresh wampee fruit.

[0014] Specifically, in step (1), when making pulp from yellow peel, the seeds of yellow peel are removed first (seedless yellow peel does not need to be removed), and then pulp is made. Furthermore, this invention does not impose any special restrictions on the pulping process.

[0015] Preferably, the mass ratio of pectinase, cellulase and hemicellulase is (2.6-3.4):(0.08-1.2):1.

[0016] More preferably, the mass ratio of pectinase, cellulase and hemicellulase is (2.8-3.2):(0.09-1.1):1.

[0017] More preferably, the mass ratio of pectinase, cellulase, and hemicellulase is 3:1:1.

[0018] Preferably, in step (1), the pressure of the first ultra-high pressure treatment is 90-330 MPa; the time of the first ultra-high pressure treatment is 1.5-6 min.

[0019] More preferably, in step (1), the pressure of the first ultra-high pressure treatment is 100-300 MPa; the time of the first ultra-high pressure treatment is 2-5 min.

[0020] Preferably, in step (1), the temperature of the enzymatic hydrolysis is 40-60℃, and the time of the enzymatic hydrolysis is 55-100 min.

[0021] More preferably, in step (1), the temperature of the enzymatic hydrolysis is 45-55℃; and the time of the enzymatic hydrolysis is 60-90 min.

[0022] Preferably, in step (2), the mass ratio of the enzymatic hydrolysis of yellow peel pulp to the extraction solvent is 1:0.8-2.2.

[0023] More preferably, in step (2), the mass ratio of the enzymatic hydrolysis of yellow peel pulp to the extraction solvent is 1:1-2.

[0024] Preferably, in step (2), the extraction solvent is selected from at least one of pure water, methanol, and ethanol.

[0025] Preferably, in step (2), the alkaline solution is selected from at least one of sodium hydroxide and potassium hydroxide.

[0026] Specifically, the main function of adding alkali solution is twofold: firstly, to adjust the pH; and secondly, because the yellow peel contains some flavonoids, which are cross-linked with polysaccharides, lignin, and other components to form insoluble bound flavonoids that are difficult to extract. Under the action of alkali solution and ultra-high pressure, these insoluble flavonoids can be transformed into soluble substances, thereby hydrolyzing the insoluble bound flavonoids and dissolving them, thus enriching the composition of flavonoid compounds.

[0027] Preferably, after adding the alkaline solution, the pH is adjusted to 10-14; more preferably, the pH is adjusted to 11-13.

[0028] Preferably, in step (2), the pressure of the second ultra-high pressure treatment is 350-600 MPa; the time of the second ultra-high pressure treatment is 4-12 min.

[0029] More preferably, in step (2), the pressure of the second ultra-high pressure treatment is 400-600 MPa; the time of the second ultra-high pressure treatment is 5-10 min.

[0030] Specifically, the extraction time is the time required for the second ultra-high pressure treatment.

[0031] Preferably, in step (3), the volume ratio of the yellow peel pulp to the solvent is 1:0.8-2.2.

[0032] More preferably, in step (3), the volume ratio of the yellow peel pulp to the solvent is 1:1-2.

[0033] Preferably, in step (3), the solvent is selected from at least one of methanol and ethanol.

[0034] Specifically, the flavonoids extracted in step (3) are prone to precipitation and separation from other substances under these conditions. Adding methanol and / or ethanol can prevent the loss caused by precipitation and separation. In addition, microorganisms are prone to grow in water systems. Adding methanol and / or ethanol can also have a certain antiseptic effect.

[0035] Preferably, the acid solution is selected from at least one of hydrochloric acid, citric acid, and malic acid.

[0036] Specifically, flavonoids are unstable under alkaline conditions but relatively stable under acidic conditions. Adding acid to adjust the pH of the system can improve the stability of flavonoids.

[0037] Preferably, the pH is 2.5-5.5; more preferably, the pH is 3.0-5.0.

[0038] Preferably, in step (3), the flavonoids include at least four of the following: gentic acid, caffeic acid, dihydroquercetin, kaempferol-7-glucoside, o-coumaric acid, rutin, quercetin-3-glucoside, norihesperidin, and hesperidin.

[0039] A second aspect of the present invention provides a flavonoid.

[0040] Specifically, the flavonoids are extracted by the method for extracting flavonoids from the peel of the plant described in the first aspect of the present invention; the flavonoids include at least four of the following: gentian acid, caffeic acid, dihydroquercetin, kaempferol-7-glucoside, o-coumaric acid, rutin, quercetin-3-glucoside, norihesperidin, and tangeretin.

[0041] Compared with the prior art, the beneficial effects of the technical solution provided by the present invention are as follows:

[0042] (1) This invention uses a combination of enzymatic method, ultra-high pressure treatment and alkaline method to extract flavonoids from yellow peel. This extraction method can not only improve the extraction efficiency of flavonoids, but also promote the conversion of insoluble bound flavonoids into free flavonoids, which can greatly improve the extraction yield of flavonoids, enrich the composition of flavonoid compounds, and has good application value.

[0043] (2) The complex enzyme can hydrolyze pectin and cellulose in wampee, reducing their impact on the extraction of flavonoids. The first ultra-high pressure treatment not only activates the activity of the complex enzyme but also ensures sufficient contact between the complex enzyme and the pectin and cellulose in wampee, improving the efficiency of enzymatic hydrolysis. The first ultra-high pressure treatment also alters the structure of pectin, making it easier to decompose. The second ultra-high pressure treatment ensures sufficient contact between the extraction solvent and the wampee pulp, increasing the flavonoid extraction yield and shortening the extraction time.

[0044] (3) By adding alkali solution to adjust the pH of the system to make it alkaline, the insoluble bound flavonoids can be hydrolyzed and dissolved. The combined action of alkali and the second ultra-high pressure can improve the dissolution rate, thereby increasing the extraction yield of flavonoids. Detailed Implementation

[0045] To enable those skilled in the art to more clearly understand the technical solutions described in this invention, the following embodiments are provided for illustration. It should be noted that the following embodiments do not constitute a limitation on the scope of protection claimed by this invention.

[0046] Unless otherwise specified, the raw materials, reagents or devices used in the following examples are available from conventional commercial sources or can be obtained by existing known methods.

[0047] Example 1

[0048] A method for extracting flavonoids from yellow peel includes the following steps:

[0049] (1) Take fresh wampee, remove the seeds, and pulp. Take 100g of wampee pulp and add 0.006% pectinase, 0.002% cellulase, and 0.002% hemicellulase according to the weight of the wampee pulp. Mix well and then perform the first ultra-high pressure treatment at a pressure of 200MPa for 2min. Then, enzymatically hydrolyze the wampee pulp in a water bath at 45℃ for 90min to obtain enzymatically hydrolyzed wampee pulp.

[0050] (2) Take the enzymatic hydrolysis of yellow peel pulp obtained in step (1), add pure water at a mass ratio of 1:1, adjust the pH to 13 with 20% sodium hydroxide solution, and carry out a second ultra-high pressure treatment at a pressure of 500 MPa for 5 min to obtain yellow peel pulp treated with two ultra-high pressures.

[0051] (3) Take the yellow peel pulp obtained in step (2) after secondary ultra-high pressure treatment, add methanol at a mass ratio of 1:1, then add hydrochloric acid to adjust the pH to 3.0, filter with filter cloth to obtain the extract of flavonoids.

[0052] Example 2

[0053] A method for extracting flavonoids from yellow peel includes the following steps:

[0054] (1) Take fresh wampee, remove the seeds, and pulp it. Take 100g of wampee pulp and add 0.012% pectinase, 0.004% cellulase, and 0.004% hemicellulase according to the weight of the wampee pulp. Mix well and then perform the first ultra-high pressure treatment at a pressure of 300MPa for 3min. Then, enzymatically hydrolyze it in a water bath at 55℃ for 90min to obtain enzymatically hydrolyzed wampee pulp.

[0055] (2) Take the enzymatic hydrolysis of yellow peel pulp obtained in step (1), add pure water at a mass ratio of 1:2, adjust the pH to 12 with 20% sodium hydroxide solution, and carry out a second ultra-high pressure treatment at a pressure of 550 MPa for 8 min to obtain yellow peel pulp treated with two ultra-high pressures.

[0056] (3) Take the yellow peel pulp obtained in step (2) after secondary ultra-high pressure treatment, add methanol at a mass ratio of 1:1, then add hydrochloric acid to adjust the pH to 5.0, filter with filter cloth to obtain the extract of flavonoids.

[0057] Example 3

[0058] A method for extracting flavonoids from yellow peel includes the following steps:

[0059] (1) Take fresh wampee, remove the seeds, and pulp it. Take 100g of wampee pulp and add 0.03% pectinase, 0.01% cellulase, and 0.01% hemicellulase according to the weight of the wampee pulp. Mix well and then perform the first ultra-high pressure treatment at a pressure of 250MPa for 5min. Then, enzymatically hydrolyze it in a water bath at 50℃ for 80min to obtain enzymatically hydrolyzed wampee pulp.

[0060] (2) Take the enzymatic hydrolyzed yellow peel pulp obtained in step (1), add pure water at a mass ratio of 1:2, adjust the pH to 11 with 20% sodium hydroxide solution, and carry out a second ultra-high pressure treatment at a pressure of 550 MPa for 10 min to obtain yellow peel pulp treated with two ultra-high pressures.

[0061] (3) Take the yellow peel pulp obtained in step (2) after secondary ultra-high pressure treatment, add methanol at a mass ratio of 1:1, then add hydrochloric acid to adjust the pH to 4.5, filter with filter cloth to obtain the extract of flavonoids.

[0062] Comparative Example 1

[0063] Comparative Example 1 is a commonly used alcohol extraction method in the prior art.

[0064] A method for extracting flavonoids from yellow peel includes the following steps:

[0065] (1) Take fresh yellow peel, remove the seeds, and pulp. Take 100g of yellow peel pulp and add 0.03% pectinase, 0.01% cellulase and 0.01% hemicellulase according to the weight of yellow peel pulp. Mix well and then enzymatically hydrolyze in a 50℃ water bath for 80min to obtain enzymatically hydrolyzed yellow peel pulp.

[0066] (2) Take the enzymatic hydrolyzed yellow peel pulp obtained in step (1), add ethanol at a mass ratio of 1:2, extract for 180 min, filter with filter cloth to obtain the extract of flavonoids.

[0067] Comparative Example 2

[0068] The difference between Comparative Example 2 and Example 3 is that in step (1), no compound enzyme was added to Comparative Example 2 and no enzymatic hydrolysis was performed, while the rest was the same as in Example 3.

[0069] Comparative Example 3

[0070] The difference between Comparative Example 3 and Example 3 is that in step (1), Comparative Example 3 did not undergo the first ultra-high pressure treatment, while the rest was the same as Example 3.

[0071] Comparative Example 4

[0072] The difference between Comparative Example 4 and Example 3 is that in step (2), Comparative Example 4 did not undergo a second ultra-high pressure treatment, and the extraction time was 10 min, while the rest was the same as Example 3.

[0073] Comparative Example 5

[0074] The difference between Comparative Example 5 and Example 3 is that in step (2), no alkali solution was added to Comparative Example 5, while the rest was the same as in Example 3.

[0075] Performance testing

[0076] The extracts from Examples 1-3 and Comparative Examples 1-5 were diluted to 1000 mL with methanol. The total flavonoid content, flavonoid composition, and scavenging abilities of 2,2-diphenyl-1-picrylhydrazine (DPPH) and 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt cation (ABTS) were then determined. +Free radical scavenging capacity, iron ion reducing capacity, and oxygen radical absorbance capacity (ORAC). Total flavonoid content and antioxidant capacity (DPPH scavenging capacity, ABTS scavenging capacity) in the extracts of Examples 1-3 and Comparative Examples 1-5. + The free radical capacity, iron ion reducing capacity, and ORAC are shown in Table 1; the composition of flavonoids in the extracts of Examples 1-3 and Comparative Examples 1-5 is shown in Table 2.

[0077] 1. Determination of total flavonoid content

[0078] The total flavonoid content in the extracts of Examples 1-3 and Comparative Examples 1-5 was determined using the sodium nitrite-aluminum nitrate method.

[0079] Take 6 mL of the extract diluted with methanol, add 1 mL of 5% NaNO2 solution, shake to mix, let stand for 5 min, then add 1 mL of 10% Al(NO3)3 solution, shake to mix, let stand for 6 min, then add 4 mL of 1 mol / L NaOH solution, shake to mix, place in a 45℃ water bath for 10 min, remove and cool, then centrifuge at 1000 rpm for 5 min, and measure the absorbance of the supernatant at a wavelength of 505 nm. Rutin was used as the standard sample. The total flavonoid content in the extracts of Examples 1-3 and Comparative Examples 1-5 is shown in Table 1.

[0080] 2. Determination of the composition of flavonoids

[0081] The composition of flavonoids in the extracts of Examples 1-3 and Comparative Examples 1-5 was determined by liquid chromatography-mass spectrometry (LC-MS).

[0082] Chromatographic conditions: C18 column (150 mm × 4.6 mm, 2.6 μm); mobile phase A: acetonitrile (containing 0.2% formic acid by volume); mobile phase B: 0.6% formic acid solution by volume. Gradient elution program: mobile phase A initially at 0% volume, increased to 10% after 10 min, increased to 15% after 30 min, held for 10 min, increased to 20% after 50 min, increased to 30% after 60 min and held for 5 min, decreased to 5% after 66 min and held for 4 min.

[0083] The injection volume was 10 μL, the detection wavelengths were 280 and 330 nm, the flow rate was 0.600 mL / min, and the column temperature was maintained at 40 °C.

[0084] MS conditions: electrospray ionization source, positive and negative ion scanning mode, capillary voltage 4.5kV; nebulizer pressure 1.5bar; drying temperature 300℃, drying gas flow rate 9.0L / min, mass scan range 100-1000m / z.

[0085] The composition of flavonoids in the extracts of Examples 1-3 and Comparative Examples 1-5 is shown in Table 2.

[0086] 3. Determination of DPPH free radical scavenging ability

[0087] Take 50 μL of the diluted extracts from Examples 1-3 and Comparative Examples 1-5, add 150 μL of DPPH solution (0.2 mmol / L), mix well, and react in the dark at room temperature for 20 min. Measure the absorbance at 517 nm using a microplate reader. Using water-soluble vitamin E (Trolox) as a standard, determine the DPPH free radical scavenging rate of different mass concentrations of Trolox and plot a standard curve. The experiment also included a reagent blank group (anhydrous ethanol), a control group (using an equal volume of methanol instead of the sample), and a sample blank group (using an equal volume of anhydrous ethanol instead of the DPPH solution). The DPPH free radical scavenging ability of the sample is expressed as Trolox equivalents. The DPPH scavenging rate is calculated using the following formula.

[0088]

[0089] In the formula: A1 is the absorbance of the control group; A0 is the absorbance of the reagent blank group; A i A represents the absorbance of the sample group. j This represents the absorbance of the blank sample group.

[0090] The DPPH free radical scavenging ability of the extracts from Examples 1-3 and Comparative Examples 1-5 is shown in Table 1.

[0091] 4. Clear ABTS + Free radical capacity measurement

[0092] Mix 50 mL of 7 mmol / L ABTS solution and 0.88 mL of 140 mmol / L potassium persulfate solution, and let stand overnight at room temperature in the dark to prepare ABTS stock solution. Before use, dilute with anhydrous ethanol to an absorbance of 0.7 ± 0.02 at 734 nm.

[0093] Take 10 μL of the diluted extracts from Examples 1-3 and Comparative Examples 1-5, add 200 μL of ABTS solution, vortex for 30 s, and react for 6 min at room temperature in the dark. Measure the absorbance at 734 nm using a microplate reader. Using Trolox as a standard, determine the effect of different mass concentrations of Trolox on ABTS. + To determine the free radical scavenging rate, a standard curve was plotted. The experiment included a reagent blank group, a control group, and a sample blank group. The scavenging rate was calculated using the following formula:

[0094]

[0095] In the formula: A1 is the absorbance of the control group; A0 is the absorbance of the reagent blank group; A i A represents the absorbance of the sample group. j This represents the absorbance of the blank sample group.

[0096] ABTS Removal with Extracts from Examples 1-3 and Comparative Examples 1-5 + The free radical capacity is shown in Table 1.

[0097] 5. Determination of iron ion reducing power

[0098] Take 1 mL of the diluted extracts from Examples 1-3 and Comparative Examples 1-5, add 0.2 mL of 0.2 mol / L phosphate buffer (pH 6.6) and 1.5 mL of 0.3% potassium ferricyanide, mix well, incubate at 50°C for 20 min, then rapidly cool and add 1 mL of 10% trichloroacetic acid, mix well, centrifuge at 3000 r / min for 10 min, take 2 mL of the supernatant, add 0.5 mL of 0.3% ferric chloride solution, mix well, then add 3 mL of pure water, shake well, and measure the absorbance at a wavelength of 700 nm. Using Trolox as a standard, the iron reducing power of the sample is expressed as the Trolox equivalent (μmol / g). The iron ion reducing power of the extracts from Examples 1-3 and Comparative Examples 1-5 is shown in Table 1.

[0099] 6. ORAC determination

[0100] The extracts from Examples 1-3 and Comparative Examples 1-5 were diluted with 75 mmol / L phosphate buffer at pH 7.4 (v / v: 1:10) for later use.

[0101] Add 20 μL of diluted sample to each well of a black 96-well plate and incubate at 37°C. Add 80 μL of 1.25 μmol / L sodium fluorescein solution (prepared with 75 mmol / L phosphate buffer at pH 7.4) and incubate at 37°C for 5 min. Add 100 μL of 140 mmol / L 2,2'-azobis-2-methyl-propanimidamide dihydrochloride (AAPH) to each well, vortex to mix, and measure the fluorescence intensity of each well using a microplate reader. Use 120 μL of distilled water mixed with 80 μL of 1.25 μmol / L sodium fluorescein solution as a negative control, and 20 μL of distilled water mixed with 80 μL of 1.25 μmol / L sodium fluorescein solution and 100 μL of 140 mmol / L AAPH as a blank control. 500 μmol / L Trolox 4, 8, 12, 16 and 20 μL were respectively transferred into 96-well plates, and distilled water was added to 20 μL to prepare a standard curve.

[0102] Fluorescence measurement conditions: excitation wavelength 485 nm; emission wavelength 520 nm, 35 cycles, 2.5 min per cycle. Trolox was used as a standard, and the ORAC value of the sample was expressed as Trolox equivalent (μmol / g).

[0103] The ORAC extracts from Examples 1-3 and Comparative Examples 1-5 are shown in Table 1.

[0104] Table 1: Total flavonoid content and antioxidant capacity in extracts from Examples 1-3 and Comparative Examples 1-5

[0105]

[0106] As shown in Table 1, the total flavonoid content in the extracts of Examples 1-3 is higher than that in the extracts of Comparative Examples 1-5, especially higher than that obtained by alcohol extraction in Comparative Example 1 using existing technology. The differences between Comparative Examples 2-5 and Example 3 are that no compound enzyme was added, no first ultra-high pressure treatment was performed, no second ultra-high pressure treatment was performed, and no alkali treatment was performed, respectively. This resulted in significantly lower total flavonoid content and antioxidant capacity in the extracts of Comparative Examples 2-5 compared to Example 3. This indicates that the extraction method of the present invention, which combines enzymatic method, first ultra-high pressure treatment, second ultra-high pressure treatment, and alkali treatment, can improve the extraction yield of flavonoids, and the extracted extract has good antioxidant capacity.

[0107] Table 2: Composition of flavonoids in the extracts of Examples 1-3 and Comparative Examples 1-5

[0108]

[0109]

[0110] As shown in Table 2, the extracts of Examples 1-3 of this invention contain a wide variety of flavonoids, including gentic acid, caffeic acid, dihydroquercetin, kaempferol-7-glucoside, o-coumaric acid, rutin, quercetin-3-glucoside, norihesperidin, and hesperidin. The extracts of Comparative Example 1 contain gentic acid, caffeic acid, o-coumaric acid, and rutin; the extracts of Comparative Example 2 contain gentic acid, caffeic acid, o-coumaric acid, rutin, and norihesperidin; the extracts of Comparative Example 3 contain gentic acid, caffeic acid, o-coumaric acid, rutin, norihesperidin, and hesperidin; the extracts of Comparative Example 4 contain gentic acid, caffeic acid, o-coumaric acid, rutin, quercetin-3-glucoside, and norihesperidin; and the extracts of Comparative Example 5 contain gentic acid, caffeic acid, o-coumaric acid, and rutin. Among them, Comparative Examples 1 and 5 showed the fewest types of flavonoids, while Examples 1-3 showed the most types. Dihydroquercetin and kaempferol-7-glucoside were only detected in Examples 1-3. This indicates that the extract obtained using the extraction method of the present invention contains a wide variety of flavonoids, which can enrich the composition of flavonoids and has good application value.

[0111] The above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A method for extracting flavonoids from the skin of a fruit, characterized by, Includes the following steps: (1) Pulp the yellow peel, add compound enzyme, and then perform a first ultra-high pressure treatment and enzymatic hydrolysis to obtain enzymatic hydrolyzed yellow peel pulp; (2) Add the enzymatic hydrolyzed yellow peel pulp obtained in step (1) to the extraction solvent, then add alkali solution, and then perform a second ultra-high pressure treatment to obtain yellow peel pulp treated with two ultra-high pressures. (3) Add the solvent to the yellow peel pulp obtained in step (2), then add acid, and separate the solid and liquid to obtain flavonoids; In step (1), the complex enzyme is composed of pectinase, cellulase, and hemicellulase; In step (2), the pressure of the second ultra-high pressure treatment is 350-600 MPa; the time of the second ultra-high pressure treatment is 4-12 min. In step (3), the volume ratio of the yellow peel pulp to the solvent is 1:0.8-2.2; the solvent is selected from at least one of methanol and ethanol; The flavonoids include gentian acid, caffeic acid, dihydroquercetin, kaempferol-7-glucoside, o-coumaric acid, rutin, quercetin-3-glucoside, norihesperidin, and hesperidin.

2. The extraction method according to claim 1, characterized in that, In the complex enzyme, the mass ratio of pectinase, cellulase, and hemicellulase is (2.6-3.4):(0.08-1.2):

1.

3. The extraction method according to claim 1, characterized in that, In step (1), the pressure of the first ultra-high pressure treatment is 90-330 MPa; the time of the first ultra-high pressure treatment is 1.5-6 min.

4. The extraction method according to claim 1, characterized in that, In step (1), the temperature of the enzymatic hydrolysis is 40-60℃; the time of the enzymatic hydrolysis is 55-100 min.

5. The extraction method according to claim 1, characterized in that, In step (2), the mass ratio of the enzymatic hydrolysis of yellow peel pulp to the extraction solvent is 1:0.8-2.

2.

6. The extraction method according to claim 5, characterized in that, In step (2), the extraction solvent is selected from at least one of pure water, methanol, and ethanol.

7. The extraction method according to claim 1, characterized in that, In step (2), the alkaline solution is selected from at least one of sodium hydroxide and potassium hydroxide.