Method for extracting polyphenol from persimmon peel and application

Abstract

The invention provides a method for extracting polyphenol from persimmon peel and application, the method comprises the steps of persimmon peel pretreatment, irradiation treatment, subcritical ethanol extraction and extraction of polyphenol in different forms, and the method belongs to the technical field of natural product extraction methods. According to the method, electron beam irradiation is utilized to treat persimmon peels, high energy in electron beams can bombard components such as cellulose and lignin in cell walls of the persimmon peels, tightly-combined chemical bonds are broken, ethanol in a subcritical extraction technology can permeate into the persimmon peels to extract polyphenol substances, and therefore the extraction efficiency of polyphenol is improved; and the consumption of the solvent in the extraction process can be reduced. Different forms of polyphenols are further extracted on the basis of the persimmon peel polyphenol crude extract, the polyphenol has good antioxidant capacity and alpha-glucosidase inhibitory activity, bioactive compounds in persimmon peel can be efficiently recycled, and future waste utilization and development of natural antioxidants are promoted.

Description

Technical Field

[0001] The invention belongs to the technical field of natural product extraction methods, and specifically relates to a method for extracting polyphenols from persimmon peel and application thereof. Background Art

[0002] Persimmon trees belong to the Diospyros family and are popular worldwide for their unique taste and rich nutritional value. In 2021, China's persimmon production reached 3.4 million tons, accounting for nearly 75% of the global total production. Dried persimmon fruit (persimmon cake) is the main processed product of persimmon. It is worth noting that at least 10% of persimmon peel is produced as a by-product during the production process, and these by-products are usually treated as waste. The polyphenols in persimmon play a key role in alleviating or preventing oxidative stress and are expected to be used as a natural medicine for anti-aging, blood sugar control and metabolic health. In addition, studies have shown that the polyphenol content in persimmon peel is higher than that in the pulp and has stronger antioxidant activity, so persimmon peel may become a promising new source of polyphenols. Extraction techniques have a significant impact on polyphenol content. Studies have shown that microwave-assisted extraction of phenolic compounds from Brazilian olive leaves increased the total phenol content (TPC) by 1.82 times, and the application of ultrasound-assisted extraction in the extraction of phenolic compounds from waste coffee grounds found that TPC increased by up to 50%. Subcritical fluid extraction is increasingly seen as a viable alternative to traditional methods because it can improve extraction efficiency while reducing the use of toxic chemicals and energy and shortening extraction time. In addition, subcritical solvent extraction can effectively extract polar compounds by adjusting the dielectric constant by changing temperature and pressure. Based on laboratory-scale research results, subcritical ethanol extraction has been applied to the extraction of phenolic compounds on a pilot scale. At the same time, under the conditions of 120°C and 0.5MPa, using a batch reactor with a volume scale-up factor of 1000, the total yields of phenolic compounds and flavonoids in buckwheat waste reached 29.8g / kg and 13.9g / kg, respectively. Therefore, subcritical-assisted extraction is expected to become an effective technology for the extraction and utilization of polyphenols from persimmon peel. Many studies have shown that the combined use of new extraction technologies can bring significant synergistic benefits by increasing the extraction rate of target compounds while reducing the consumption of organic solvents. For example, electron beam irradiation (EBI) can destroy cell walls and improve the extraction efficiency of phenolic compounds. Studies have reported that the application of EBI pretreatment to the extraction of active substances in walnut green peel increased the total phenol content by 1.18 times, flavonoids by 1.43 times, and triterpenes by 1.11 times. Therefore, electron beam irradiation coupled with subcritical extraction technology has great potential in the efficient recovery of bioactive compounds in persimmon peel, which can promote future waste utilization and the development of natural antioxidants. Summary of the invention

[0003] Technical problems to be solved: In view of the above technical problems, the purpose of the present invention is to provide a method for extracting polyphenols from persimmon peel, the steps of which include persimmon peel pretreatment, irradiation treatment, subcritical ethanol extraction, and extraction of polyphenols in different forms, belonging to the technical field of natural product extraction methods. The present invention utilizes electron beam irradiation to treat persimmon peel. The high energy in the electron beam can bombard the cellulose, lignin and other components in the cell wall of the persimmon peel, so that the tightly bound chemical bonds are broken, which is conducive to the ethanol infiltration of the subcritical extraction technology into the persimmon peel to extract polyphenols, thereby improving the extraction efficiency of polyphenols and reducing the consumption of solvents in the extraction process. The present invention further extracts polyphenols in different forms based on the crude extract of persimmon peel polyphenols, has good antioxidant capacity and α-glucosidase inhibitory activity, can efficiently recover bioactive compounds in persimmon peel, and promotes future waste utilization and the development of natural antioxidants.

[0004] Technical solution: A method for extracting polyphenols from persimmon peel, comprising the following steps: S1. Crush and sieve the persimmon peel to make persimmon peel powder; S2. The persimmon peel powder was subjected to electron beam irradiation to obtain irradiated persimmon peel powder; S3. Add ethanol solution to the irradiated persimmon peel powder and mix well, perform subcritical ethanol extraction to obtain persimmon peel polyphenols and solid residue. Furthermore, the conditions for the irradiation treatment in step S2 are an irradiation dose of 10-20 kGy and an irradiation time of 5-10 s. Furthermore, the concentration of the ethanol solution in step S3 is 70-80%. Furthermore, in step S3, the solid-liquid ratio of the irradiated persimmon peel powder to the ethanol solution is 1:(40-50). Furthermore, the conditions for subcritical ethanol extraction in step S3 are extraction temperature 140-160° C. and extraction time 20-40 min. Application of persimmon peel polyphenols extracted by any of the above methods in the development of antioxidants and α-glucosidase inhibitors. Application of persimmon peel polyphenols extracted by any of the above methods in the preparation of products for improving blood sugar levels and anti-oxidation. Furthermore, the method also includes extracting insoluble persimmon peel polyphenols from the solid residue. Furthermore, the extraction method of the insoluble persimmon peel polyphenols is as follows: ① adding the solid residue to a sodium hydroxide solution containing EDTA and ascorbic acid for hydrolysis at room temperature for 4 hours, adjusting the pH of the solution to 2, and then repeatedly extracting with ethyl acetate to obtain insoluble ester-bound phenol and aqueous phase residue; ② the aqueous phase residue is acid-hydrolyzed with hydrochloric acid at 75°C for 60 minutes, adjusting the pH of the solution to 2, and then repeatedly extracting with ethyl acetate to obtain insoluble glycosidic bond-bound phenol. Use of the insoluble persimmon peel polyphenols extracted by any of the methods described above in the preparation of products for improving blood sugar levels. Beneficial effects: 1. The present invention utilizes an electron beam irradiation-subcritical ethanol extraction method to extract polyphenols from persimmon peels. The high-energy electron beams generated by the irradiation bombard the cell walls of the persimmon peels to generate a large number of free radicals. The free radicals attack the chemical bonds of components such as cellulose, hemicellulose and pectin in the cell walls of the persimmon peels, causing them to break and the cell wall structure to become loose and easy to decompose, which is beneficial to the penetration of ethanol solvents and the dissolution of polyphenols in subcritical ethanol extraction, thereby improving the efficiency of subcritical ethanol extraction of polyphenols. 2. The persimmon peel polyphenols extracted in the present invention have excellent antioxidant properties and can significantly inhibit the activity of DPPH free radicals and hydroxyl free radicals. At the same time, the persimmon peel polyphenols are separated into different forms of phenolic substances to obtain free phenol (FP), soluble ester-bound phenol (SEBP), soluble glycosidic-bound phenol (SGBP), insoluble ester-bound phenol (ISEBP) and insoluble glycosidic-bound phenol (ISGBP), which have good α-glucosidase inhibitory activity. 3. The persimmon peel polyphenol extraction method provided by the present invention can not only broaden the research direction of persimmon peel polyphenol extraction, but also provide technical support for the extraction of biologically active substances in persimmon peel in the future, increase the product added value of persimmon peel, promote the high-value utilization of persimmon peel, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 is the total phenol content in the persimmon peel extracts of Example 1 and Comparative Examples 1-4; Figure 2 The antioxidant capacity of Example 1 and Comparative Example 1; Figure 3 The α-glucosidase inhibitory activity of FP, SEBP, SGBP, ISEBP and ISGBP in the persimmon peel polyphenols of Example 1 (EBI-SAE) and Comparative Example 1 (CE) was evaluated. DETAILED DESCRIPTION The present invention will be further described below in conjunction with embodiments, which are explanations of the present invention and are not limited to the following embodiments: Example 1 A method for extracting persimmon peel polyphenols by electron beam irradiation-subcritical ethanol extraction comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2.50g of persimmon peel powder was irradiated with electron beam for 7.5s at a irradiation dose of 15kGy to obtain irradiated persimmon peel powder; S3. Add 2.5 L of 70% ethanol solution to the irradiated persimmon peel powder and mix well, place in a subcritical extraction device, and extract at 160° C. for 40 min to obtain persimmon peel polyphenols and solid residue. Example 2 A method for extracting persimmon peel polyphenols by electron beam irradiation-subcritical ethanol extraction comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2.50g of persimmon peel powder was irradiated with electron beam for 5s at a irradiation dose of 10kGy to obtain irradiated persimmon peel powder; S3. Add 2.5 L of 70% ethanol solution to the irradiated persimmon peel powder and mix well, place in a subcritical extraction device, and extract at 160° C. for 40 min to obtain persimmon peel polyphenols and solid residue. Example 3 A method for extracting persimmon peel polyphenols by electron beam irradiation-subcritical ethanol extraction comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2.50g of persimmon peel powder was irradiated with electron beam for 10s at a irradiation dose of 20kGy to obtain irradiated persimmon peel powder; S3. Add 2.5 L of 70% ethanol solution to the irradiated persimmon peel powder and mix well, place in a subcritical extraction device, and extract at 160° C. for 40 min to obtain persimmon peel polyphenols and solid residue. Example 4 A method for extracting persimmon peel polyphenols by electron beam irradiation-subcritical ethanol extraction comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2.50g of persimmon peel powder was irradiated with electron beam for 7.5s at a irradiation dose of 15kGy to obtain irradiated persimmon peel powder; S3. Add 2.5 L of 70% ethanol solution to 50 g of persimmon peel powder and mix well. Place in a subcritical extraction device and extract at 120° C. for 40 min to obtain persimmon peel polyphenols and solid residue. Example 5 A method for extracting persimmon peel polyphenols by electron beam irradiation-subcritical ethanol extraction comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2.50g of persimmon peel powder was irradiated with electron beam for 7.5s at a irradiation dose of 15kGy to obtain irradiated persimmon peel powder; S3. Add 2.5 L of 70% ethanol solution to 50 g of persimmon peel powder and mix well. Place in a subcritical extraction device and extract at 140° C. for 40 min to obtain persimmon peel polyphenols and solid residue. Comparative Example 1 The difference between this comparative example and Example 1 is that the traditional solvent method is adopted. A method for extracting persimmon peel polyphenols using a traditional solvent comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2. Add 1.25 L of 70% ethanol solution to 50 g of persimmon peel powder and mix well. Place the mixture in an extraction device and extract at 40° C. for 40 min to obtain persimmon peel polyphenols and a solid residue. Comparative Example 2 The difference between this comparative example and Example 1 is that subcritical extraction is not used. A method for extracting persimmon peel polyphenols using electron beam irradiation-traditional solvents comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2.50g of persimmon peel powder was treated under the condition of irradiation dose of 15kGy to obtain irradiated persimmon peel powder; S3. Add 1.25 L of 70% ethanol solution to the irradiated persimmon peel powder, mix well, place in an extraction device, and extract at 40° C. for 40 min to obtain persimmon peel polyphenols and solid residue. Comparative Example 3 The difference between this comparative example and Example 1 is that an ultrasonic-assisted step is added, and electron beam irradiation and subcritical extraction are not used. A method for extracting persimmon peel polyphenols using ultrasonic treatment-traditional solvents comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2. Add 1.25 L of 70% ethanol solution to 50 g of persimmon peel powder and mix well. Place the mixture in an extraction device and extract for 40 min at an ultrasonic power of 450 W and 40° C. to obtain persimmon peel polyphenols and a solid residue. Comparative Example 4 The difference between this comparative example and Example 1 is that an ultrasonic-assisted step is added and subcritical extraction is not used. A method for extracting persimmon peel polyphenols by electron beam irradiation-ultrasonic treatment-traditional solvent comprises the following steps: S1.100g persimmon peel was crushed and sieved to obtain persimmon peel powder; S2.50g of persimmon peel powder was treated under the condition of irradiation dose of 15kGy to obtain irradiated persimmon peel powder; S2. Add 1.25 L of 70% ethanol solution to the irradiated persimmon peel powder and mix well. Place the mixture in an extraction device and extract for 40 min at an ultrasonic power of 300 W and 40° C. to obtain persimmon peel polyphenols and a solid residue. Example 6 The method for extracting insoluble persimmon peel polyphenols from the solid residue in Example 1 comprises the following steps: S1. Extraction of insoluble ester-bound phenol (ISEBP) The solid residue was reacted with a 4 mol / L NaOH solution containing 10 mmol / L EDTA and 1% ascorbic acid at room temperature in a ratio of 1:20 (w / v) to obtain the ISEBP component. After the reaction was continued for 4 h, the pH value of the suspension was adjusted to 2, and then the suspension was mixed with an ethyl acetate solution in a ratio of 1:1 (v / v) to separate the ISEBP component; S2. Extraction of Insoluble Glycosidically-Bound Phenol (ISGBP) The remaining aqueous phase residue was hydrolyzed with 6 mol / L HCl at 75°C for 60 min, and then the pH value of the suspension was adjusted to 2. ISGBP was extracted from the acidified supernatant with ethyl acetate (1:1, v / v). Example 7 The method for extracting insoluble persimmon peel polyphenols from the solid residue in Example 1 comprises the following steps: S1. Extraction of insoluble ester-bound phenol (ISEBP) The solid residue was reacted with a 4 mol / L NaOH solution containing 10 mmol / L EDTA and 1% ascorbic acid at room temperature in a ratio of 1:15 (w / v) to obtain the ISEBP component. After the reaction lasted for 3 h, the pH value of the suspension was adjusted to 2, and then the suspension was mixed with an ethyl acetate solution in a ratio of 1:1 (v / v) to separate the ISEBP component; S2. Extraction of Insoluble Glycosidically-Bound Phenol (ISGBP) The remaining aqueous phase residue was hydrolyzed with 6 mol / L HCl at 75°C for 60 min, and then the pH value of the suspension was adjusted to 2. ISGBP was extracted from the acidified supernatant with ethyl acetate (1:1, v / v). Example 8 The method for extracting insoluble persimmon peel polyphenols from the solid residue in Example 1 comprises the following steps: S1. Extraction of insoluble ester-bound phenol (ISEBP) The solid residue was reacted with a 4 mol / L NaOH solution containing 10 mmol / L EDTA and 1% ascorbic acid at room temperature in a ratio of 1:20 (w / v) to obtain the ISEBP component. After the reaction lasted for 3 h, the pH value of the suspension was adjusted to 2, and then the suspension was mixed with an ethyl acetate solution in a ratio of 1:1 (v / v) to separate the ISEBP component; S2. Extraction of Insoluble Glycosidically-Bound Phenol (ISGBP) The remaining aqueous phase residue was hydrolyzed with 6 mol / L HCl at 60°C for 90 min, and then the pH value of the suspension was adjusted to 2. ISGBP was extracted from the acidified supernatant with ethyl acetate (1:1, v / v). Comparative Example 5 The difference between this comparative example and Example 4 is that the solid residue is obtained by the method of Comparative Example 1. The method for extracting insoluble persimmon peel polyphenols from the solid residue in Comparative Example 1 comprises the following steps: S1. Extraction of insoluble ester-bound phenol (ISEBP) The solid residue was reacted with a 4 mol / L NaOH solution containing 10 mmol / L EDTA and 1% ascorbic acid at room temperature in a ratio of 1:20 (w / v) to obtain an ISEBP component. After the reaction was continued for 4 hours, the pH value of the suspension was adjusted to 2, and then the suspension was mixed with an ethyl acetate solution in a ratio of 1:1 (v / v) to separate the ISEBP component; S2. Extraction of Insoluble Glycosidically-Bound Phenol (ISGBP) The remaining aqueous phase residue was hydrolyzed with 6 mol / L HCl at 75°C for 60 min, and then the pH value of the suspension was adjusted to 2. ISGBP was extracted from the acidified supernatant with ethyl acetate (1:1, v / v). Performance Testing (1) Total phenolic content (TPC) TPC was determined using the Folin-Ciocalteau reagent method. Phenolic components (500 μL) were mixed with 2.5 mL of Folin-Ciocalteau reagent (10%, v / v) and sodium carbonate solution (2 mL, 7.5%, w / w). After reacting at room temperature in the dark for 30 minutes, the absorbance was measured at 765 nm and expressed as milligrams of gallic acid per gram of raw material (dry weight) (mg GAE / g DW), wherein Example 1 was recorded as electron beam-subcritical ethanol, Comparative Example 1 was recorded as traditional solvent, Comparative Example 2 was recorded as electron beam-solvent, Comparative Example 3 was recorded as ultrasound-solvent, Comparative Example 4 was recorded as electron beam-ultrasound-solvent, and Comparative Example 5 was recorded as subcritical ethanol. Table 1 Total phenol content of Examples 1-3 and Comparative Examples 1-5 Total phenolic content (mgGAE / gDW) Example 1 31.94±0.54 Example 2 28.35±0.83 Example 3 28.03±0.37 Example 4 12.21±0.72 Example 5 21.76±0.21 Comparative Example 1 7.10±0.12 Comparative Example 2 8.04±0.06 Comparative Example 3 7.14±0.18 Comparative Example 4 5.68±0.03 From Table 1 and Figure 1 It can be seen that the total phenol content of Example 1 is significantly higher than that of Comparative Examples 1-4, indicating that subcritical ethanol extraction can allow ethanol solvent molecules to quickly diffuse into the persimmon peel under a subcritical state and fully contact with the polyphenols in the persimmon peel. In addition, the ethanol mass transfer rate under the subcritical state is fast, which can accelerate the dissolution of polyphenols from the persimmon peel, thereby ensuring the high efficiency of persimmon peel extraction. The synergistic effect of electron beam irradiation can first utilize the high energy effect of the electron beam to disintegrate the structure in the persimmon peel, making it loose and easy to disperse, and further accelerate the penetration of the ethanol solvent. (2) Extraction rate of insoluble persimmon peel polyphenols The contents of insoluble ester-bound phenol (ISEBP) and insoluble glycosidic-bound phenol (ISGBP) in Example 6, Example 7, Example 8 and Comparative Example 5 were determined. Table 2 Insoluble persimmon peel polyphenol content of Example 4, Example 5, Example 6 and Comparative Example 5 As can be seen from Table 2, Examples 4-6 were all separated from the solid residue of Example 1, while Comparative Example 5 was separated from the solid residue of Comparative Example 1. The contents of insoluble ester-bound phenol (ISEBP) and insoluble glycosidic-bound phenol (ISGBP) in Examples 4-6 were lower than those in Comparative Example 5. This is because the high-energy electron beam generated by electron beam irradiation can destroy the chemical bonds in the insoluble polyphenols in persimmon peel and convert them into soluble polyphenols, which are then extracted during the subcritical ethanol extraction process, resulting in a decrease in the content of insoluble polyphenols in the remaining solid residue. (3) Total flavonoid content (TFC) TFC was determined using the sodium nitrite-aluminum chloride method with slight modifications. 200 μL of sample was placed in a 1.5 mL centrifuge tube, and then 550 μL of sodium nitrite solution (5%, w / w), 75 μL of aluminum chloride solution (10%, w / w) and 500 μL of sodium hydroxide solution (1 mol / L) were added in sequence, with reaction times of 5, 6 and 15 min, respectively. The results were measured at 510 nm and expressed as milligrams of rutin per gram of raw material (dry weight) (mg RE / g DW). Table 3 Soluble total phenol and total flavonoid contents of Example 1, Comparative Example 1 and Comparative Example 5 Total phenols (mgGAE / gDW) Total flavonoids (mgRE / gDW) Comparative Example 1 7.10±0.14 5.11±0.09 Example 1 31.94±0.54 24.53±2.38 As can be seen from Table 3, the total phenol content and total flavonoids content of Example 1 are higher than those of Comparative Example 1, while the total sugar content is lower than that of Comparative Example 1, indicating that electron beam irradiation in the electron beam irradiation-subcritical ethanol extraction method can make the persimmon peel structure loose, and the diffusion rate and mass transfer rate of ethanol in the subcritical state are accelerated, which can accelerate the dissolution of bioactive substances in the persimmon peel in ethanol, so the total phenol content and total flavonoids content increase. (5) Evaluation of antioxidant capacity ① DPPH free radical scavenging ability determination: accurately draw 100 μL of sample, then add 1 mL of DPPH working solution (concentration of 50 μg / L in methanol), react at room temperature and in the dark for 30 minutes, and then measure the absorbance at 517 nm. The result is expressed as milligrams of VC equivalent per milliliter of extract (mgAAE / g DW). ② ABTS free radical scavenging ability determination: ABTS·+ stock solution was obtained by mixing 7mM ABTS and 2.45mM K2S2O8 and incubating for 16h at room temperature in the dark. The obtained ABTS·+ solution was then diluted with 80% (v / v) ethanol to prepare the working reagent, and the pH was maintained at 0.7±0.02. 100μL of sample was mixed with 2mL ABTS working solution. After reacting for 6min at room temperature in the dark, the absorbance was measured at 734nm, and the results were expressed as mgAAE / g DW. ③ Determination of hydroxyl radical (·OH) scavenging ability: 1 mL salicylic acid solution (9 mmol / L in ethanol), 1 mL diluted sample, 1 mL FeSO 4 (9mmol / L) and 1mLH 2 O 2 After mixing with 0.01%, the mixture was reacted at 37°C for 15 min, and the absorbance was recorded at a wavelength of 510 nm. The result was expressed as mgAAE / g DW. Table 4 Evaluation of the antioxidant capacity of Example 1 and Comparative Example 1 From Table 4 and Figure 2 It can be seen that the DPPH free radical scavenging ability, ABTS free radical scavenging ability and hydroxyl free radical scavenging ability of Example 1 are all higher than those of Comparative Example 1, indicating that the persimmon peel polyphenols extracted by electron beam irradiation-subcritical ethanol have better biological activity and better antioxidant activity. (6) Evaluation of α-glucosidase inhibition ability The persimmon peel polyphenols extracted in Example 1 and Comparative Example 1 were further separated to obtain soluble polyphenols in different forms, as follows: ① Free phenol (FP) extraction: persimmon peel polyphenols were adjusted to pH 2 with 6 mol / L hydrochloric acid, and then n-hexane (1:1, v / v) was added for defatting. Free phenol (FP) was extracted from the acidified supernatant with ethyl acetate (1:1, v / v), and the extraction was repeated 3 times; ② Extraction of soluble ester-bound phenol (SEBP): The collected organic layer was evaporated to dryness at 40°C. After removing the solvent, the phenolic components were redissolved in methanol. The aqueous solution was hydrolyzed in a 4 mol / L sodium hydroxide solution containing 10 mmol / L ethylenediaminetetraacetic acid and 1% ascorbic acid at room temperature for 4 h. The soluble ester-bound phenol (SEBP) was extracted from the acidified (pH = 2) supernatant with ethyl acetate (1:1, v / v); ③ Extraction of soluble glycosidic-bound phenol (SGBP): The remaining aqueous solution after extraction of SEBP was hydrolyzed with 6 mol / L hydrochloric acid at 75°C for 60 min, and then soluble glycosidic-bound phenol (SGBP) was extracted from the acidified (pH = 2) supernatant with ethyl acetate (1:1, v / v). The α-glucosidase inhibition rates of phenols in different forms in Example 1 and Example 4, Comparative Example 1 and Comparative Example 5 were measured respectively, as follows: 100 μL of samples of different concentrations were mixed with 100 μL of α-glucosidase (0.2 U / mL, dissolved in 0.1 mM phosphate buffer, pH 6.8), and then 50 μL of α-pNPG (2.5 mM) was added to each well as a substrate. After incubation at 37°C for 20 min, the absorbance was recorded at 405 nm. The calculation formula of inhibition rate: inhibition rate (%) = [1-(A sample -A black ) / (A control -A blank )]×100% Where A sample A represents the absorbance of samples containing inhibitors and enzymes. black A represents the absorbance of the sample without enzyme (containing buffer and enzyme). control represents the absorbance of the control sample containing buffer and enzyme, A blank The results are expressed as the sample concentration required to inhibit the enzyme activity by 50% (IC 50 ) was expressed in milligrams per milliliter (mg / mL) and determined by linear regression analysis. Depend on Figure 3 It can be seen that EBI-SAE represents electron beam irradiation-subcritical ethanol extraction (Example 1), CE represents traditional solvent extraction (Comparative Example 1), and the α-glucosidase IC of different forms of phenols in Example 1 and Example 4 50The size order is FP < ISEBP < SEBP < ISGBP < SGBP, with the range being 0.14 - 1.85 mg / mL. For Comparative Example 1 and Comparative Example 5, the α-glucosidase IC of different forms of phenols 50 The size order is ISEBP < FP < ISGBP < SGBP < SGEP, with the range being 0.15 - 6.47 mg / mL. Additionally, in Example 1 and Comparative Example 1, FP and ISEBP have lower α-glucosidase IC 50 , showing good α-glucosidase inhibitory ability. At the same time, the α-glucosidase IC of FP, SGBP, SGEP, and ISGBP in Example 1 and Example 4 50 are all lower than those in Comparative Example 1 and Comparative Example 5, indicating that compared with traditional solvent extraction, different forms of phenolic substances in persimmon peel polyphenols extracted by electron beam irradiation-subcritical ethanol have more excellent α-glucosidase inhibitory activity. Additionally, the α-glucosidase IC of ISEBP in Example 4 50 is higher than that in Comparative Example 5 because, under the same volume condition, the content of ISEBP in Comparative Example 5 is about 4 times that of ISEBP in Example 4, and its inhibitory activity against α-glucosidase is strong, so its α-glucosidase IC 50 is lower than that in Example 4. The above are only the preferred embodiments of the present invention and do not impose any formal restrictions on the present invention. Any person skilled in the art can make many possible changes and modifications to the technical solution of the present invention, or modify it into an equivalent embodiment with equivalent changes, without departing from the spirit and technical solution of the present invention. Therefore, any simple modification, equivalent replacement, equivalent change, and modification made to the above embodiments based on the technical essence of the present invention without departing from the technical solution of the present invention still fall within the scope of protection of the technical solution of the present invention.

Claims

1. A method for extracting polyphenols from persimmon peel, characterized in that: The following steps are involved: S1. Crush and sieve the persimmon peel to make persimmon peel powder; S2. The persimmon peel powder was subjected to electron beam irradiation to obtain irradiated persimmon peel powder; S3. Add ethanol solution to the irradiated persimmon peel powder and mix well, perform subcritical ethanol extraction to obtain persimmon peel polyphenols and solid residue.

2. The method for extracting polyphenols from persimmon peel according to claim 1, characterized in that: The conditions of the irradiation treatment in step S2 are an irradiation dose of 10-20 kGy and an irradiation time of 5-10 s.

3. The method for extracting polyphenols from persimmon peel according to claim 1, characterized in that: The concentration of the ethanol solution in step S3 is 70-80%.

4. The method for extracting polyphenols from persimmon peel according to claim 1, characterized in that: In the step S3, the solid-liquid ratio of the irradiated persimmon peel powder to the ethanol solution is 1:(40-50).

5. The method for extracting polyphenols from persimmon peel according to claim 1, characterized in that: The conditions for the subcritical ethanol extraction in step S3 are extraction temperature 140-160° C. and extraction time 20-40 min.

6. Use of persimmon peel polyphenols extracted according to the method according to any one of claims 1 to 5 in developing antioxidants and α-glucosidase inhibitors.

7. Use of persimmon peel polyphenols extracted according to the method described in any one of claims 1 to 5 in preparing products for improving blood sugar levels and anti-oxidation.

8. The method for extracting polyphenols from persimmon peel according to claim 1, characterized in that: The method also includes extracting insoluble persimmon peel polyphenols from the solid residue.

9. The method for extracting polyphenols from persimmon peel according to claim 8, characterized in that: The insoluble persimmon peel polyphenol extraction method is as follows: ① adding the solid residue to a sodium hydroxide solution containing EDTA and ascorbic acid for hydrolysis at room temperature for 4 hours, adjusting the solution pH to 2, and then repeatedly extracting with ethyl acetate to obtain insoluble ester-bonded phenol and aqueous phase residue; ② The aqueous phase residue was hydrolyzed with hydrochloric acid at 75°C for 60 min, the pH of the solution was adjusted to 2, and then repeatedly extracted with ethyl acetate to obtain insoluble glycosidic bond-bound phenol.

10. Use of the insoluble persimmon peel polyphenols extracted by the method according to any one of claims 8 to 9 in preparing products for improving blood sugar levels.

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