Avocado seed polyphenol and preparation method thereof, and avocado seed starch and preparation method thereof

Abstract

The invention relates to the technical field of plant polysaccharide and polyphenol extraction, in particular to avocado seed polyphenol and a preparation method thereof, and avocado seed starch and a preparation method thereof. The scheme of the invention provides a method for synchronously extracting polyphenol and starch from avocado seeds by using an oxidative coupling supercritical CO2 extraction technology, and the method specifically comprises the following steps: peeling the avocado seeds, crushing, continuously drying and crushing after spontaneous oxidation reaction of the avocado seeds, and primarily extracting by using an extracting agent; further extracting polyphenol by using a supercritical CO2 fluid extraction device; and finally, freeze-drying polyphenol obtained by two times of extraction to obtain polyphenol solid, and adding a proper amount of protease into residual insoluble substances for deproteinization treatment to obtain starch. The starch prepared by adopting the scheme is low in digestibility, the prepared polyphenol has good thermal stability, and the prepared polyphenol and the polyphenol-starch compound have good tyrosinase inhibitor activity and antioxidant activity.

Description

Technical Field

[0001] The present application relates to the technical field of plant polysaccharide and polyphenol extraction, and in particular to an avocado seed polyphenol and a preparation method thereof, an avocado seed starch and a preparation method thereof. Background Art

[0002] Avocado (Persea americana Mill.), also known as alligator and avocado, belongs to the genus Persea genus of the Lauraceae family. It is native to Mexico and Central America and is mostly planted in tropical and subtropical regions of the world. Avocado is rich in plant fat, so it is known as the "forest cream". Specifically, the fat content in avocado flesh is around 12-31%, of which unsaturated fatty acids account for the largest proportion. The main fatty acids are oleic acid (50-60%), followed by saturated palmitic acid (15-20%), unsaturated palmitic acid (6-10%), and polyunsaturated linoleic acid (11-15%). In addition to being rich in polyunsaturated fatty acids, avocados have high nutritional value and functional ingredients that are beneficial to human health, such as carotenoids, dietary fiber, potassium, magnesium, and vitamins. Vitamin E, plant sterols, squalene, etc., and low in sugar, it is a high-energy, low-sugar fruit, so it is classified as a functional food. It is reported that in addition to being sold as an edible fruit, avocado is also an important raw material for humans to obtain high-value vegetable oils. There are many literatures focusing on the effects of different extraction methods on the quality of oils extracted from avocado pulp. The results show that the oils extracted from avocado pulp are not only edible, but also widely used in the cosmetics industry to produce various skin or hair care products because of its low acidity, non-irritating to the skin, and ability to promote transdermal absorption.

[0003] Looking at human consumption habits, humans only eat the flesh of avocados, and often discard the peel and kernel (i.e., seeds) as "waste" in the environment. The industry also crushes the peel or kernel and uses it as low-cost waste for composting or animal feed. However, previous reports have shown that avocado peel or kernel contains rich carbohydrates, small amounts of lipids and proteins. Obviously, the current processing method has caused a serious waste of avocado by-product resources.

[0004] Therefore, the biggest practical problem facing the avocado industry is that its by-products have not been utilized in a high-value manner, which is not conducive to the healthy development of the avocado industry. Based on this, it is meaningful to explore how to efficiently utilize the beneficial components in avocado by-products.

[0005] Among them, on the one hand, starch is an important substance that provides energy for human biological activities. It is the main component of human staple foods such as bread, steamed bread, rice, etc. It is particularly important for human health development, so the market demand is huge. At present, edible starch mainly comes from cereals or potato crops, so can starch be extracted from avocado seeds? It is meaningful to explore whether starch can be extracted from avocado by-products and how to effectively extract starch, so as to recycle avocado by-products.

[0006] On the other hand, colorants play an important role in improving the appearance and quality of food. Synthetic pigments with cost advantages can give food and beverage products stable and uniform colors. However, with the strict supervision of laws and regulations in various countries and the increasing health awareness of more and more consumers, the importance of natural pigments in the food industry has become increasingly prominent. At present, the main natural plant pigments are carotenoids, chlorophyll, betalain and anthocyanins. The roots, stems, leaves, flowers and fruits of plants with rich colors (such as orange peel, pumpkin, dragon fruit, tomato, etc.) are important sources of natural pigments (mainly polyphenols). So can polyphenol pigments be extracted from avocado seeds? It is meaningful to explore whether it is possible to extract natural pigments with thermal stability from avocado by-products and how to effectively extract natural pigments, which can reuse avocado by-products and increase the added value of avocado.

[0007] Those skilled in the art know that seeking starch or polyphenol pigments from different sources is conducive to developing new food resources and enriching the source channels of starch or polyphenols for the food industry. However, no relevant reports on the simultaneous extraction of starch, polyphenol pigments, etc. from avocado seeds have been found so far.

[0008] In summary, avocado is a high-quality fruit with rich nutritional value and many benefits to the human body. It is an important raw material for extracting functional lipids. However, there has been no relevant report on the extraction of starch, polyphenol pigments and other substances from avocado seeds alone (the raw material is only avocado seeds, without pulp and peel), especially the extraction of polyphenol substances with good active functions from avocado seeds alone. At present, the utilization rate of its processing by-products (avocado seeds) is low, resulting in serious waste of resources. Summary of the invention

[0009] In order to solve the problems mentioned in the prior art mentioned in the above background technology, this application provides the following technical solutions: The present application provides a method for preparing avocado seed polyphenols, which comprises the following steps: Pretreatment: remove the outer skin of avocado seeds and crush them into particles; place the particles for spontaneous oxidation reaction (0 to 60 minutes); after the spontaneous oxidation reaction is completed, dry the particles, crush them and sieve them; The first step of extraction: adding an extractant to the dried particles, stirring the particles throughout the process and performing ultrasonic treatment at intervals for a certain period of time for 1 to 2 hours, filtering to obtain a polyphenol supernatant and a first solid, drying the first solid and then ultrafine grinding to obtain avocado seed powder; The second step is extraction: the avocado seed powder is loaded into an extraction column for supercritical CO2 fluid extraction. After dynamic extraction for 1 to 3 hours, the polyphenol extract and the second solid are recovered by separation treatment; the polyphenol supernatant and the polyphenol extract are mixed and dried to obtain avocado seed polyphenols; wherein, the supercritical treatment temperature is adjusted to 30 to 40°C and the supercritical treatment pressure is adjusted to 20 to 30 MPa so that the extraction fluid is in a supercritical state, and the flow rate of the entrainer is 0.2 to 0.6 mL / min.

[0010] In some embodiments, the extractant is one of 75% anhydrous ethanol, 85% anhydrous ethanol, 95% anhydrous ethanol, anhydrous ethanol, ethyl acetate, and acetone; the entrainer is one of 75% anhydrous ethanol, 85% anhydrous ethanol, 95% anhydrous ethanol, anhydrous ethanol, ethyl acetate, and acetone.

[0011] In some embodiments, the extractant is 85% anhydrous ethanol; the entrainer is 85% anhydrous ethanol.

[0012] In some embodiments, the extractant is ethyl acetate and the entrainer is ethyl acetate.

[0013] In some embodiments, in the pretreatment step, the outer skin of the avocado seeds is first removed and crushed into particles with a particle size of less than 0.2 cm; the particles are placed in an aerobic environment at 25 to 40°C for 0 to 60 minutes for a spontaneous oxidation reaction; after the spontaneous oxidation reaction is completed, the particles are dried to a moisture content of between 10 and 15%, and crushed through an 80-mesh sieve.

[0014] In some embodiments, in the first extraction step, an extractant is added to the particles at a mass ratio of particles to extractant of 1:(15-20), and then the particles are stirred throughout the process, and ultrasonic treatment is performed for (2-5) minutes every (5-15) minutes, with an ultrasonic power of 400W-600W for 1-2 hours. The polyphenol supernatant and the first solid are filtered out, and the first solid is dried and then ultrafinely crushed to obtain avocado seed powder.

[0015] In some embodiments, in the second extraction step, avocado seed powder is loaded into an extraction column, and then the extraction column is connected to the gas pipeline of a supercritical CO2 extractor, the supercritical treatment temperature is adjusted to 30-40°C and the supercritical treatment pressure is adjusted to 20-30MPa, so that the extraction fluid is in a supercritical state, the flow rate of the entrainer is adjusted to (0.2-0.6) mL / min, and the dynamic extraction is performed for 1-3 hours; after the extraction is completed, the polyphenol extract and the second solid are recovered by vacuum separation; the polyphenol supernatant and the polyphenol extract are then mixed and freeze-dried to obtain avocado seed polyphenols.

[0016] The present application provides a method for preparing avocado seed starch, which comprises the following steps: preparing starch by enzymatic hydrolysis: mixing a second solid with water, adding bromelain and papain, and stirring for a certain period of time; then separating and treating to recover the starch solid, and washing and drying the starch solid to obtain avocado seed starch; wherein the second solid is prepared by the method for preparing avocado seed polyphenols as described above; and the amount of bromelain and the amount of papain used are both 0.5% to 1% of the mass of the second solid.

[0017] In some embodiments, in the enzymatic starch production step, the second solid is mixed with water in a mass ratio of 1: (10-15), and then bromelain and papain are added, and stirred for 1h-2h; then centrifuged to recover the starch solids, and the starch solids are washed and dried to obtain avocado seed starch.

[0018] The present application also provides an avocado seed polyphenol, which is prepared using the avocado seed polyphenol preparation method as described above.

[0019] The present application also provides an avocado seed starch, which is prepared using the avocado seed starch preparation method as described above.

[0020] Based on the above, compared with the prior art, the preparation method of avocado seed polyphenols provided in this application has the following technical principles and effects: The present application utilizes oxidation reaction coupled with supercritical CO2 fluid extraction technology to extract polyphenols and polyphenol-starch complexes from avocado seeds. The prepared polyphenols and polyphenol-starch complexes have good thermal stability, which can be used as natural plant-extracted pigments, and the prepared polyphenols and polyphenol-starch complexes have good tyrosinase inhibitor activity and antioxidant activity. Therefore, the present application fully reuses the discarded seeds of avocados, avoids the waste of food resources, opens up new food sources for seeking polyphenols and other raw materials from different sources, and enriches the selection channels of polyphenols and other raw materials in the food industry.

[0021] Other features and beneficial effects of the present application will be described in the following description, and partly become apparent from the description, or understood by practicing the present application. The purpose and other beneficial effects of the present application can be realized and obtained by the contents shown in the description, claims and drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a physical visual picture of the products avocado seed polyphenols, polyphenol-starch complex, and starch obtained in Example 5; Figure 2 The microstructure of avocado seed polyphenols and starch obtained in Example 5; Figure 3 The X-ray photoelectron spectrum of avocado seed polyphenols and starch obtained in Example 5; Figure 4 The infrared spectra and X-ray diffraction patterns of avocado seed polyphenols, starch, and polyphenol-starch complexes obtained in Example 5; Figure 5 This is the NMR spectrum of the polyphenol-starch complex obtained in Example 5; Figure 6 This is the NMR spectrum of the avocado seed polyphenols obtained in Example 2; Figure 7 This is the NMR spectrum of the starch product obtained in Example 5; Figure 8 The hydroxyl radical scavenging ability data of the products obtained in different embodiments and comparative example 7 are shown in FIG. Fig. 9 ABTS of the products obtained in different embodiments and comparative example 7 + Free radical scavenging rate data graph; Fig.10 This is a physical picture of the product polyphenol obtained in Comparative Example 7. DETAILED DESCRIPTION

[0023] In order to make the purpose, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, not all of the embodiments; the technical features designed in different implementation modes of the present application described below can be combined with each other as long as they do not conflict with each other; based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of this application.

[0024] In the description of this application, it should be noted that all terms (including technical terms and scientific terms) used in this application have the same meanings as those generally understood by ordinary technicians in the field to which this application belongs, and cannot be understood as limitations on this application; it should be further understood that the terms used in this application should be understood to have meanings consistent with the meanings of these terms in the context of this specification and in the relevant fields, and should not be understood in an idealized or overly formal sense, unless explicitly defined in this application.

[0025] 1. The present application provides the following examples and comparative examples to verify the effects of the present application scheme: Example 1 The preparation steps are as follows: Step (1), pre-treatment of avocado seeds: The outer skin of avocado seeds is removed and crushed into small particles with a particle size of less than 0.2 cm; the particles are placed under aerobic conditions at 30°C for 45 minutes. After the spontaneous oxidation reaction is completed, the discolored avocado seed particles are dried to a moisture content of 12% and crushed through an 80-mesh sieve.

[0026] Step (2), the first step of extracting avocado seed polyphenols: The extractant was added at a mass ratio of 1:15 between avocado seed particles and the extractant, and then the extraction process was stirred throughout, and ultrasonic waves were applied for 5 minutes every 15 minutes at a power of 500 W. After 2 hours, the polyphenol supernatant was filtered and collected for later use. The filtered lower solid was dried and further ultra-finely crushed to obtain avocado seed powder (i.e., polyphenol-starch complex).

[0027] Wherein, the extractant of Example 1 is 75% anhydrous ethanol.

[0028] Step (3), second step extraction of avocado seed polyphenols: Weigh an appropriate amount of the ultrafinely crushed avocado seed powder (polyphenol-starch complex) obtained by the first step of extraction, load it into a stainless steel extraction column, connect the extraction column to the gas pipeline of the supercritical CO2 fluid extractor, adjust the temperature to 35°C and the pressure to 25MPa, so that the extraction fluid is in a supercritical state, adjust the entrainer to set the flow rate to 0.3mL / min, and perform dynamic extraction for 1h, 2h, and 3h respectively. After the extraction, separate under reduced pressure to recover the avocado seed polyphenol extract and avocado seed powder solid matter.

[0029] The polyphenol supernatant obtained by the first step of extraction and the avocado seed polyphenol extract obtained by the second step of extraction are mixed, and freeze-dried to obtain avocado seed polyphenols. The freeze-drying cold trap temperature is -80°C, and the drying is carried out under vacuum conditions for 24 hours.

[0030] Wherein, the entrainer in Example 1 is 75% anhydrous ethanol.

[0031] Step (4): Enzymatic starch production According to the mass ratio of avocado seed powder (i.e., polyphenol-starch complex) to water of 1:12, water was added to the avocado seed powder, and then 0.75% of the solid matter mass of the avocado seed powder obtained after the second step of extraction and separation treatment, 0.5% of the solid matter mass of papain (enzyme activity 30,000 U / g) was added, and stirred for 1 hour. The solid was collected by centrifugation, washed with water 3 times, and dried to obtain avocado seed starch without protein.

[0032] Example 2-6 (Based on Example 1, the types of extractant and entrainer were changed) The difference between Example 2-6 and Example 1 is that the extractants and entrainers used are different. The extractants of Examples 2-6 are 85% anhydrous ethanol, 95% anhydrous ethanol, anhydrous ethanol, ethyl acetate, and acetone, respectively, and the entrainers are 85% anhydrous ethanol, 95% anhydrous ethanol, anhydrous ethanol, ethyl acetate, and acetone, respectively. The other preparation processes and conditions are the same as those of Example 1.

[0033] Example 7-10 (Based on Example 3, the spontaneous oxidation time was changed) The difference between Examples 7-10 and Example 3 is that the spontaneous oxidation time is different, which is 0 min, 15 min, 30 min, and 60 min in Example 7-10. The other preparation processes and conditions are the same as those in Example 3.

[0034] Comparative Example 1-6 (lack of the first step of extraction) Comparative Examples 1-6 correspond to the case where the first step of extraction in Examples 1-6 is omitted, and the seed powder pre-treated in step (1) is directly applied to the supercritical CO2 fluid extraction in step (3).

[0035] Moreover, three experimental groups with supercritical pressures of 20, 25, and 30 MPa were respectively conducted in each comparative example.

[0036] Comparative Example 7 (method described in Chinese invention patent with patent application number 202010472437.0) Preparation of avocado powder: Take the peel and core of avocado, clean them, dry them in a 50°C oven, crush them, sieve them to obtain avocado powder, mix them with an extractant (using 60wt% ethanol solution as solvent, 0.1% citric acid monoglyceride + 0.01% uridine diphosphate glucose + 60wt% ethanol solution), place them in an ultrasonic device (ultrasonic power 40KHz or 60KHz) for extraction for 2h, cool them, centrifuge them, and collect the filtrate; Purification of polyphenols: Take the pretreated macroporous resin, add the filtrate, elute with 70wt% ethanol solution, statically adsorb for 6h, collect the eluate, concentrate to remove alcohol, and vacuum freeze-dry at a temperature of -5°C, a vacuum pressure of 50Pa, and a time of 1h. Polyphenols are obtained.

[0037] Among them, the preparation method of the pretreated macroporous resin is: seal and soak the AB-8 macroporous resin in 95wt% ethanol for 24 hours, wash with distilled water until there is no ethanol smell, then soak it in 5% sodium hydroxide solution for 12 hours, wash it with distilled water until it is neutral and has no turbidity, and finally soak it in 5% hydrochloric acid solution for 12 hours, wash it with distilled water until it is neutral and has no turbidity.

[0038] In Comparative Example 7, two parallel experimental groups were conducted at ultrasonic powers of 40 KHz and 60 KHz, namely, Comparative Example 7 Experimental Group 1 and Comparative Example 7 Experimental Group 2, respectively.

[0039] 2. Implementation Example Test Data Analysis 1. Exploring the effect of the type of extractant on the first step polyphenol extraction rate in Examples 1-6 The extraction rate of polyphenols obtained by the first step of extraction in Examples 1-6 was measured, as shown in Table 1 below: The extraction rate test process is as follows: take 1 mL of the polyphenol supernatant obtained in the first step of extraction and put it into a -80°C refrigerator for rapid freezing to solid form, and then put it into a freeze dryer for drying. After drying, the first step polyphenol powder is obtained and weighed. The ratio of the weight of the first step polyphenol powder obtained after freeze-drying the extract after the first step of extraction to the volume of the extract is recorded as the extraction rate of the first step polyphenol.

[0040] Table 1 Effect of solvent type on the first step polyphenol extraction rate

[0041] Result analysis: In order to improve the extraction efficiency of plant pigments (the polyphenols extracted in this application can be used as plant extraction pigments), a suitable extraction method must be selected. Generally, plant pigments are extracted using traditional methods, such as Soxhlet extraction, solid phase extraction, and steam distillation. Traditional extraction methods are simple, economical, and easy to use, but there are problems such as solvent residues and time consumption. Water, ethanol, and methanol are most commonly used to extract polar and water-soluble pigments, and non-polar solvents such as hexane, acetone, trifluoroethylene and other organic solvents are used to extract lipophilic pigments.

[0042] However, due to the complex composition of avocado seed polyphenols, there are currently no reports on the extraction of polyphenol pigments therefrom, so it is not yet known which reagent is more conducive to extracting more polyphenols.

[0043] The effects of different extractants on the extraction rate of avocado seed polyphenols are shown in the table above. The data show that: This application scheme uses 75% ethanol, 85% ethanol, 95% ethanol, anhydrous ethanol, ethyl acetate, and acetone as extraction agents, and it is found that they all have good extraction effects. The results show that 85% ethanol is more suitable as an extraction agent for avocado seed polyphenols. The reason for the analysis is that the reason why 85% ethanol is used as an extraction agent is mainly affected by the ratio of water / fat-soluble polyphenol content in avocado seeds, and 85% ethanol is more compatible with the components in avocado seeds.

[0044] 2. Exploring the effect of entrainer type on the second step polyphenol extraction rate in Examples 1-6 The extraction rate of the second-step polyphenols obtained by the second-step extraction in Examples 1-6 was measured, as shown in the following Table 2: The extraction rate test process is as follows: take 1 mL of avocado seed polyphenol extract obtained in the second step of extraction, put it into a -80°C refrigerator and quickly freeze it into a solid to obtain the second step polyphenol powder, put it into a freeze dryer, and after it is completely dried, the second step polyphenol powder is obtained and weighed. The ratio of the weight of the second step polyphenol powder obtained after freeze-drying the extract after the second step of extraction to the volume of the extract is recorded as the extraction rate of the second step polyphenol.

[0045] Table 2 Effect of entrainer type on the extraction rate of polyphenols in the second step

[0046] Analysis results: The use of supercritical fluid extraction technology in the food industry should be one of the most promising processes. In the food and pharmaceutical industries, supercritical CO2 is used as a solvent. It has the advantages of being non-toxic, harmless, and will not cause pollution to the extracted material. It solves the problems of solvent residue in organic solvent extraction methods. The process is simple, and only the main parameters such as temperature and pressure need to be controlled to achieve the purpose of extracting different components in the mixture; the extraction rate is high and the selectivity is good; the extraction temperature is low, and it has a good protective effect on heat-sensitive and easily oxidized components, and can well maintain the activity of the extracted material; the extraction solvent can be recycled and the production cost is low. A small amount of components with strong affinity to the extracted material are added to the supercritical fluid to improve its selectivity and solubility to the extracted material. This type of substance added is called an entrainer.

[0047] However, due to the complex composition of avocado seed polyphenols, there are currently no reports on the use of supercritical fluid extraction technology to extract avocado seed polyphenols, and there are no reports on the use of the first step extraction combined with the second step supercritical fluid extraction technology. Therefore, it is not yet known which reagent is more conducive to supercritical fluid extraction of more polyphenols.

[0048] The effects of different entrainers on the extraction rate of avocado seed polyphenols are shown in the table above. The test results show that: This application scheme uses 75% ethanol, 85% ethanol, 95% ethanol, anhydrous ethanol, ethyl acetate, and acetone as entrainers, and it is found that they all have good extraction effects. The results show that 85% ethanol as an entrainer is more conducive to the extraction of more avocado seed polyphenols. The reason for the analysis is that the introduction of the entrainer increases the polarity of the solvent, thereby increasing the solubility of polar substances. In essence, the solubility of the solvent depends on the size of the molecular force between the entrainer and the polar solute. The polarity of 85% ethanol is more compatible with the components in avocado seeds.

[0049] 3. Exploring the effect of spontaneous oxidation time after seed peeling and crushing on the first step extraction rate in Examples 7-10 The extraction rate of the first step polyphenol powder obtained by the first step extraction in Examples 7-10 was measured, as shown in Table 3 below: Table 3 Data table of the effect of spontaneous oxidation time on the first step extraction rate after seed peeling and crushing

[0050] From the data in the above table, we can see that: The oxidation time after seed crushing has a significant impact on the first step of extraction efficiency. When extraction is performed immediately after crushing (0min), the polyphenols inside the seeds have not fully completed the structural transformation and have not yet formed pigment polyphenols with stable color, resulting in a low extraction rate. As oxidation forms, the oxygen in the air causes the seed polyphenols to undergo oxidative browning and complete the structural transformation. When the spontaneous oxidation time approaches 45min, the pigment inside is basically in an oxidative saturation state, completing the formation of polyphenols with structural unit temperature and showing a bright color.

[0051] Therefore, the preferred spontaneous oxidation time of the preparation method of the present application is 45 minutes, at which time the polyphenols inside the seeds fully complete the structural transformation of pigment polyphenols, resulting in a high extraction rate. Continuing oxidation after 45 minutes is also feasible, but compared with 45 minutes, the extension of time leads to a decrease in production efficiency and a small increase in extraction rate.

[0052] 4. Functional comparison 1: Exploring the effect of the selection of extractants in the examples on the tyrosinase inhibitor activity of polyphenol products: The tyrosinase inhibitor activity screening model was used to test the activity of the samples. The specific results are shown in Table 4.

[0053] The samples are: the sample in the first step of the extraction process of Example 2 (the sample obtained by freeze-drying the polyphenol supernatant in the first step, denoted as YT1), the sample in the second step of the extraction process of Example 2 (the sample obtained by freeze-drying the avocado seed polyphenol extract in the second step, denoted as CT1), the sample in the first step of the extraction process of Example 5 (the sample obtained by freeze-drying the polyphenol supernatant in the first step, denoted as YT2), and the sample in the second step of the extraction process of Example 5 (the sample obtained by freeze-drying the avocado seed polyphenol extract in the second step, CT2). Comparative Example 7: Polyphenol sample (denoted as UT1) and kojic acid prepared in Experimental Group 2.

[0054] Activity test process: 1g / L L-DOPA was used as the reaction substrate, and the tyrosinase was prepared into an enzyme solution with an enzyme activity of 100U / mL for standby use, and kojic acid was used as a positive control. 25μL samples of different mass concentrations or phosphate buffer (PBS) were added to the enzyme reaction group and the blank group, and then 25μL tyrosinase was added. After reacting at 37℃ for 5min, 100μL 1g / L L-DOPA solution was added. After reacting at 37℃ for 5min, the absorbance was measured at a wavelength of 475nm. The reaction control group and the blank control group used PBS instead of tyrosinase, and the other steps were the same as the reaction group and the blank group. Calculate the tyrosinase inhibition rate (η).

[0055] Wherein, η = [1 - (A reaction group - A reaction control) / (A blank - A blank control)] × 100%. Each sample was tested three times in parallel.

[0056] Among them, the reference of the test method is Bao Yingrong, Wang Yafeng, He Ruijie, et al. Isolation of polyphenol components from Conus foetida leaves and study on their tyrosinase inhibitory activity[J]. Chemistry and Industry of Forest Products, 2024, 44(3):112-120.

[0057] Table 4 IC values ​​of different inhibitory tyrosinase activities 50 value

[0058] Among them, IC 50 The lower the value, the better the inhibition rate of tyrosinase activity.

[0059] The experimental results show that the polyphenols obtained by supercritical CO2 extraction with ethyl acetate as the entrainer in the embodiment of the present application have a more superior effect of inhibiting tyrosinase activity, while the polyphenols obtained by extraction with aqueous ethanol solution have a relatively poor effect, indicating that the type of extractant affects the composition and structure of polyphenols, and thus affects their activity.

[0060] 5. Functional comparison 2: Exploring the effect of the selection of extractants on the antioxidant capacity of polyphenol products in the examples: Referring to the method described by Sun Yan et al. (Source: Sun Yan, Gao Shang, Guo Yuting, Chen Lu, Xue Xue, Tang Hui, Liao Bingwu, Jia Ruibo. Optimization of Ultrasonic-Assisted Extraction of Polyphenols from Elaeagnus angustifolia Leaves by Response Surface Methodology and Study on Its Antioxidant and Anti-glycation Capacity [J]. Chinese Condiments, 2024), the antioxidant capacity of the samples at different concentrations (including hydroxyl radical scavenging capacity, ABTS + Free radical scavenging rate), for detailed results, see Figure 8-9 .

[0061] The samples were: the sample during the first step of the extraction process of Example 2 (the sample obtained by freeze-drying the polyphenol supernatant in the first step, denoted as YT1), the sample during the second step of the extraction process of Example 2 (the sample obtained by freeze-drying the avocado seed polyphenol extract in the second step, denoted as CT1), the sample during the first step of the extraction process of Example 5 (the sample obtained by freeze-drying the polyphenol supernatant in the first step, denoted as YT2), and the sample during the second step of the extraction process of Example 5 (the sample obtained by freeze-drying the avocado seed polyphenol extract in the second step, CT2). The polyphenol sample (denoted as UT1) and kojic acid prepared in Experimental Group 2 of Comparative Example 7.

[0062] The experimental results show that different extraction methods (including ultrasound and supercritical CO2 fluid extraction) and the extraction reagents used have a significant impact on the antioxidant capacity of polyphenol samples. The supercritical CO2 extraction (using ethyl acetate as the extraction agent) provided in this application scheme has a superior antioxidant capacity compared to other polyphenol samples.

[0063] 6. Yield of polyphenols from avocado seeds The polyphenol supernatant obtained by the first step of extraction in Example 1-6 and the avocado seed polyphenol extract obtained by the second step of extraction were mixed and freeze-dried to obtain avocado seed polyphenols. The yield of the avocado seed polyphenols is shown in Table 5: Table 5 Yield (unit: mg of polyphenols extracted per g of avocado seeds)

[0064] The product obtained in Example 5 was subjected to the following performance tests: 1. Storage stability of avocado seed polyphenols The avocado seed polyphenols (polyphenol products obtained by combining the first step extraction and the second step extraction) and the avocado seed polyphenol-starch complex prepared in this application can both be used as pigments. Storage stability is an important property of polyphenol pigments in the industrial production process. Pigments are unstable under light irradiation and are easily decomposed, which is a property defect of many pigments. Therefore, its storage stability as a pigment is verified here: The avocado seed polyphenols and avocado seed polyphenol-starch complex, β-carotene, and sodium copper chlorophyllin prepared in Example 5 were placed under incandescent light and stored at 50°C. The retention rates of various polyphenols were tested in the first month and the third month, respectively. The retention rate test method was based on the method described by Song et al. (Source: Song, Jia, et al. "Preparation of high internal phase Pickering emulsion by centrifugation for3D printing and astaxanthin delivery." Food hydrocolloids (2023)), with slight modifications.

[0065] The specific operations are as follows: The polyphenols before storage were mixed with 4.8 ml methanol and dichloromethane (1:2, v / v). After shaking for 3 minutes and centrifuging at 5000 rpm for 5 minutes, the absorbance of the supernatant was measured at 480 nm using a Lambda 35 UV spectrophotometer (PerkinElmer, Cambridge, USA) and recorded as A1. The absorbance of the polyphenols after storage for a certain period of time was determined in the same way as A2. The measured data are shown in Table 6: Polyphenol retention rate (%) = 100 × (A2 ÷ A1); Table 6 Storage stability of polyphenols

[0066] According to the test results: like Figure 1 As shown, Figure 1 The left and middle figures are actual pictures of avocado seed polyphenols and avocado seed polyphenol-starch complexes obtained in Example 5, respectively. It can be seen from the figures that the above products have color (orange-yellow), and both can be used as natural plant-extracted pigments.

[0067] From the data in the above table, it can be seen that the avocado seed polyphenols and polyphenol-starch complexes extracted by the present application have good thermal stability, and the storage stability of avocado seed polyphenols is higher than that of β-carotene and is similar to that of sodium copper chlorophyllin. The storage stability of the avocado seed polyphenol-starch complexes extracted by the present application is higher than that of β-carotene and sodium copper chlorophyllin.

[0068] In summary, the avocado seed polyphenols and avocado seed polyphenol-starch complexes prepared by the present application have good thermal stability and can be used as natural plant-extracted pigments.

[0069] 2. Comparison of starch digestibility The nutritional value of starch depends on the proper form and amount of intake. Excessive intake of starch is associated with adverse health outcomes such as obesity, type 2 diabetes, cardiovascular disease, and hyperglycemia. Starch can be divided into four components based on its digestion rate, namely (1) rapidly digestible starch (RDS), (2) slowly digestible starch (SDS), and (3) resistant starch (RS). Foods with high SDS and RS content are considered beneficial for people with chronic and metabolic diseases because they have sugar control and other physiological functions. Here, a comparative analysis of starch digestibility of avocado seed starch prepared in this application and other daily used starches is conducted: The digestibility of different starches was determined by referring to the method of Zheng et al. (Source: Zheng Yuxue, et al. "Proanthocyanidins from Chinese berry leaves modified the physicochemical properties and digestivecharacteristic of rice starch." Food Chemistry 335(2020)).

[0070] The avocado seed starch and avocado seed polyphenol-starch complex prepared in Example 5 were used as samples, respectively, and corn starch, wheat starch, and rice starch were used as controls.

[0071] The specific determination process is as follows: 1.0 g of sample was mixed with 50 ml of sodium acetate buffer solution (pH = 5.2) and stirred in boiling water for 20 minutes to completely gelatinize the starch. Then, the sample was cooled to 37 ° C and mixed thoroughly with a simulated enzyme solution (containing 180 mg of pancreatin and 12 μL of amyloglucosidase). The mixture was stirred at 180 rpm at 37 ° C. Half a milliliter of simulated digestion solution was collected at 0 minutes, 20 minutes, and 120 minutes, and 2.5 ml of 95% methanol was added to inactivate the enzyme.

[0072] The mixed solution was centrifuged at 5000 × g (g is gravity acceleration) for 10 min, and 0.1 mL of the supernatant was mixed with 0.5 mL of invertase (7.5 mg / mL) at 37 °C to ensure that all products were converted into glucose. The glucose released from starch was measured by a glucose assay kit (GOPOD, Megazyme, Ireland). The contents of rapidly digestible starch, slowly digestible starch, and resistant starch were calculated using formulas from the literature.

[0073] The measured data are shown in Table 7: Table 7 Digestion characteristics of different starches

[0074] According to the test results: like Figure 1 As shown, Figure 1 The middle picture and the right picture are the actual pictures of the avocado seed polyphenol-starch complex and avocado seed starch prepared in Example 5, respectively. It can be seen from the pictures that the above products are powdered solids.

[0075] The digestion characteristics of different starches are shown in the table above. The results show that compared with common corn starch, wheat starch and rice starch in life, the SDS and RDS contents in avocado seed starch are higher, indicating that its digestion characteristics in the simulated gastrointestinal tract are reduced. Avocado seed starch can be suitable as a raw material for developing low-glycemic foods.

[0076] 3. Structural characterization experiment of the product of the present application example The polyphenol-starch complex, avocado seed polyphenol, and avocado seed starch obtained in Example 5 of the present application were characterized by microstructure: (1) Observation of microstructure Testing process: The sample to be tested was evenly fixed on one side of the double-sided tape, and after gold spraying, it was placed in the sample chamber of a scanning electron microscope (Pro10102, Phenom World, Holland), and the microstructure of the sample was observed at a voltage of 10 kV.

[0077] (2) Determination of surface elements and groups X-ray photoelectron spectroscopy (XPS) is an important surface analysis technology. Its working principle is to use the characteristic X-rays generated by the X-ray source to bombard the sample, stimulate photoelectrons from the sample, and introduce them into the energy analyzer. After analysis by the energy analyzer and detection by the detector, the energy distribution diagram of the photoelectrons is made, that is, the X-ray photoelectron spectrum. It can perform qualitative, quantitative and chemical state analysis of all elements except hydrogen. It is characterized by high surface sensitivity and is a micro-area and surface analysis technology.

[0078] X-ray photoelectron spectroscopy test process: The elemental composition of starch samples was measured using an X-ray photoelectron spectrometer (AXIS SUPRA+, Shimadzu, Japan) according to the method of Chen et al. (Method source: Chen, Huijing, et al. "Functionality differences between esterified and pregelatinized esterified starches simultaneously prepared by octenyl succinic anhydride modification and itsapplication in dough." International Journal of Biological Macromolecules 260(2024)). The sample to be tested was attached to a piece of tape on the sample holder. The spectrometer had been calibrated before, and the energy was referenced to C1s at 284.8 eV.

[0079] (3) Infrared spectroscopy Test process: Refer to the method of Chi et al. (Method source Chi, Hui , et al. "Synthesis of dodecenyl succinic anhydride (DDSA) corn starch." Food Research International 40.2 (2007): 232-238. FTIR analysis was performed using an infrared spectrometer (Bruker, Vertex 70, Germany). The sample to be tested was mixed with spectral grade KBr. The resolution of the Fourier transform infrared spectrum was 4 cm -1 , a total of 32 scans were performed with wave numbers ranging from 600 to 4000 cm -1 between.

[0080] (4) Nuclear magnetic resonance analysis (NMR) Test process: Referring to the method of Chi et al. (the method comes from Chi, Hui, et al. "Synthesis of dodecenyl succinic anhydride (DDSA) corn starch." Food Research International 40.2 (2007): 232-238.), a nuclear magnetic resonance spectrometer (Bruker Avance III 400, Germany) was used for structural characterization. High-resolution one-dimensional (1D) images of starch and polyphenol samples (5 mg / 0.5 mL, DMSO-d6 was used as solvent) were obtained in 256 scans. 1H spectrum.

[0081] (5) X-ray diffraction pattern (XRD) The crystal structure of the sample was determined using an X-ray diffractometer (X'Pert3 Powder10300, Malvern Panalytical, The Netherlands). The experiment was carried out under the following conditions: the diffraction pattern measurement range was 5° to 40° (2θ, step angle was 2° / min), and the target voltage and current were 40 kV and 40 mV, respectively.

[0082] Result analysis: Figure 1 The intuitive physical pictures of the polyphenol-starch complex, polyphenol, and starch respectively extracted in three stages using the scheme of the present application are shown, which are brown powdery solids.

[0083] Figure 2 The left and right pictures show the microstructure of avocado seed polyphenols and starch, respectively. Among them, polyphenols present an irregular fragmented structure. The starch particles extracted in this application are oval and smooth, which may be a typical feature of plant-derived starch. The minimum diameter of starch particles is 5 μm and the maximum diameter is about 32 μm.

[0084] like Figure 3 As shown, the element distribution on the surface of avocado seed starch and polyphenols: near 534 eV and 285 eV, both starch and polyphenols show signals corresponding to the 1s orbital electrons of O and C.

[0085] The results showed that the main structural elements of starch and polyphenols are C and O, and they are bonded in the form of C-C, C-O, and -O-C=O. In addition, avocado seed starch contains more N elements.

[0086] Figure 4 (a) shows the infrared spectra of avocado seed starch, polyphenols, and polyphenol-starch complexes, which have a peak at 765 cm -1 (CC stretch), 860~866 cm -1 (C(1)-H, CH2 deformation), 930~961 cm -1 Bands (COC glycosidic bond backbone pattern), 1018-1094 cm -1 (COH bend), 1149-1162 cm -1 (asymmetric stretching of the COC of the glycosidic bond), 1344-1348 cm -1 (COH bending, CH2 twisting) and 1415–1429 cm -1 (CH2 bending, COO stretching).

[0087] The infrared spectrum shows that the substance extracted by this scheme has characteristic peaks belonging to the basic structure of starch and polyphenols, and there are certain differences in structure among the three.

[0088] Figure 4 (b) shows the XRD patterns of avocado seed starch, polyphenols, and polyphenol-starch complexes. It is worth noting that avocado seed starch exhibits strong characteristic peaks at 15.3° and 23.5°, and unresolved double peaks at 17.1° and 18.1°, showing a typical A-type crystal structure. The starch-polyphenol complex shows typical reflections at 13.1° and 20.1°, showing a mixture of A+V crystal types, indicating the presence of starch-lipid complexes. Polyphenols present a crystal structure type that is completely different from starch.

[0089] In addition, we also use 1 H NMR spectroscopy detected the structural information of avocado seed starch and polyphenols, such as Figure 5-7 Generally speaking, 5.11–5.22 ppm are attributed to the proton peaks of carbohydrate OH-3, 2 H-1, 4.34 ppm, and 3.33–3.66 ppm are attributed to the proton peaks of carbohydrate OH-6 and H-2, 3, 4, 5, 6, respectively.

[0090] It can be seen that compared with the extracted starch, due to the more complex chemical structure of polyphenols, their hydrogen spectrum shows more proton peaks.

[0091] 3. Comparative test data analysis: 1. Explore the effect of lack of the first step of extraction on the extraction efficiency of seed polyphenols in Comparative Examples 1-6: The polyphenol extraction rates of Comparative Examples 1-6 are shown in Table 8, and the polyphenol yields are shown in Table 9: Table 8 Effects of entrainer and treatment pressure on the extraction rate of polyphenols from seed powder (without pre-extraction in the first step) extracted by supercritical CO2

[0092] Table 9 Yield

[0093] The results in the above table show that: The yield of the single extraction of comparative examples 1-6 is lower than the sum of the polyphenols extracted in the two steps of the present application. Therefore, the multi-step extraction can extract more polyphenols. The extraction pressure can affect the extraction rate of polyphenols in seed powder, but it does not always increase with the increase of pressure. When the extraction pressure is 25 MPa, the extraction rate reaches the maximum.

[0094] 2. Effects of existing methods for extracting polyphenols from avocado peel and pit: The measured data of the effect of different ultrasonic frequencies on the polyphenol extraction rate in Comparative Example 7 are shown in Table 10 below. Fig.10 As shown: Table 10 Effect of ultrasonic frequency on polyphenol extraction rate

[0095] From the above table data and Fig.10 It can be seen that: The polyphenol content obtained by the above method is relatively low, which is mainly due to the loss of polyphenols during the purification process using the macroporous resin, and the color of the product is significantly different from that of the seed polyphenols in this example, which can be seen with the naked eye.

[0096] In addition, according to the tyrosinase inhibitor activity data in Table 4 and Figure 8-9 It can be seen from the antioxidant capacity data that compared with the avocado seed polyphenol product extracted in the second step of the embodiment, the polyphenol product of Comparative Example 7 has significantly lower tyrosinase inhibition activity and poor antioxidant capacity.

[0097] In summary, the solution provided in this application has at least the following design concepts and beneficial effects: Design Concept (1) The raw material for the preparation of the present application is avocado seeds. The existing research on the extraction of avocados is focused on its pulp and peel. However, there are differences in the composition of the pulp, peel and seeds, and how this difference in composition affects the difference in their functions is still unclear. The extraction of polyphenols and starch from seeds has not been reported, resulting in a waste of resources and hindering the development of new raw materials. In addition, compared with the scheme of using avocado pulp for extraction, in addition to the raw materials coming from different parts of avocado, there are also obvious differences in the composition and content of avocado pulp and peel and the seeds selected in the present application. For example, the pulp contains a variety of vitamins, oils and proteins rich in unsaturated fatty acids, and the phenol content in the peel is higher than that of seed polyphenols. It is mainly composed of polyphenol compounds such as 5-O caffeoylquinic acid and quercetin derivatives, proanthocyanidins, etc., and cellulose, while the seeds contain polyphenol components mainly composed of flavonoids and a large amount of starch.

[0098] Therefore, although the raw materials are all derived from avocados, the different components of the raw materials themselves determine that the process and process parameters for extracting the required substances are also different. Different raw materials are combined with different preparation processes to obtain new substances with unique functions. The polyphenols obtained by the present application scheme have more superior functions of inhibiting tyrosinase activity and anti-oxidation.

[0099] (2) The present application first utilizes the spontaneous reaction of the seeds by oxidation reaction, and then performs the first step of extraction by extraction to preliminarily extract the polyphenol supernatant and obtain the starch-polyphenol extract; then the second step of extraction is performed in combination with supercritical CO2 extraction technology to extract the polyphenols from the avocado seeds. Finally, the polyphenols obtained by the two extractions are freeze-dried to obtain polyphenol solids, and an appropriate amount of protease is added to the remaining insoluble matter in the second step of extraction to perform deproteinization treatment to obtain starch.

[0100] The above extraction steps are indispensable. If there is no spontaneous oxidation reaction, some substances lack more oxygen elements in the structural unit formula, resulting in some polyphenols not completely forming structurally stable polyphenol chemicals; if there is no preliminary extraction (used to extract fat-soluble polyphenol compounds on the surface or in a free state) but a supercritical CO2 fluid extraction step is directly performed, the pressure of supercritical CO2 extraction will increase, resulting in increased costs; if only the preliminary extraction is performed without subsequent supercritical CO2 extraction, some pigments chelated inside or on the surface of carbohydrates cannot be leached, and the extraction is not sufficient, resulting in a decrease in the extraction rate of polyphenols. It can be seen that the three-step coupled extraction is used to improve the extraction rate of seed polyphenols, and the different extraction methods will result in differences in the structure of the extracted polyphenol chemicals, which will inevitably lead to differences in function or efficacy.

[0101] (3) The present application also optimizes the selection of the extraction liquid for the first step extraction and the entrainment liquid for the second step extraction. The extraction liquid for the first step extraction and the entrainment liquid for the second step extraction use 85% ethanol, which is more suitable for the extraction of components of avocado seeds and is beneficial to improving the extraction rate of polyphenols.

[0102] (4) The present application also optimizes the selection of the extracting liquid in the first step of extraction and the entraining liquid in the second step of extraction. The extracting liquid in the first step of extraction and the entraining liquid in the second step of extraction use ethyl acetate. The obtained polyphenols and polyphenol-starch complexes have more superior tyrosinase inhibitor activity and antioxidant activity.

[0103] (5) The present application also optimizes the spontaneous oxidation time, with a spontaneous oxidation time of 45 min being the best choice, thereby achieving both a high polyphenol extraction rate and high production efficiency.

[0104] (6) The present application also optimizes and designs the corresponding ultrasonic process parameters. For example, the ultrasonic power process parameter is controlled at 200w to 800w. If it is lower than the lower limit, it will cause incomplete polyphenol extraction and a large amount of residual. If it is higher than the upper limit, the extraction efficiency will tend to saturation, resulting in an adverse effect of energy waste.

[0105] Beneficial effects: The present application provides a method for extracting polyphenols, starch, and polyphenol-starch complexes from avocado seeds by utilizing an oxidation reaction coupled with supercritical CO2 extraction technology. The obtained starch has a low digestibility, and the polyphenols and polyphenol-starch complexes have good thermal stability. The obtained polyphenols and polyphenol-starch complexes have good tyrosinase inhibitor activity and antioxidant activity.

[0106] Among them, avocado seed starch can be used as a raw material for developing low-glycemic foods. Avocado seed polyphenols and avocado seed polyphenol-starch complexes have good thermal stability and can be used as natural plant-extracted pigments. The prepared polyphenols and polyphenol-starch complexes have good tyrosinase inhibitor activity and antioxidant activity, and can be added as raw material factors to functional products with corresponding effects (such as food, medicine and health products).

[0107] Therefore, the present application fully reuses discarded avocado seeds, avoids the waste of food resources, opens up new food sources for seeking starch or polyphenols from different sources, and enriches the selection channels for raw materials such as polyphenols and starch in the food and other industries.

[0108] It should be noted that: The usage of avocado seed starch includes, but is not limited to, being used as a raw material for low-glycemic foods. Similarly, the usage of avocado seed polyphenols and avocado seed polyphenol-starch complexes includes, but is not limited to, being used as natural plant-extracted pigments.

[0109] In this article, “~” is used to indicate a numerical range, and the range indicated by this expression includes two endpoint values.

[0110] In addition, those skilled in the art should understand that, although there are many problems in the prior art, each embodiment or technical solution of the present application can be improved in only one or several aspects, without having to solve all the technical problems listed in the prior art or background technology at the same time. Those skilled in the art should understand that the content not mentioned in a claim should not be used as a limitation on the claim.

[0111] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present application, rather than to limit it. Although the present application has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present application.

Claims

1. A method for preparing avocado seed polyphenols, characterized in that: The following steps are involved: Pretreatment: removing the outer skin of avocado seeds and crushing them into particles; placing the particles for spontaneous oxidation reaction for 0 to 60 minutes; after the spontaneous oxidation reaction is completed, drying, crushing and screening the particles; The first step of extraction: adding an extractant to the dried particles, stirring the particles throughout the process and performing ultrasonic treatment at intervals for a certain period of time for 1 to 2 hours, filtering to obtain a polyphenol supernatant and a first solid, drying the first solid and then ultrafine grinding to obtain avocado seed powder; The second step is extraction: the avocado seed powder is loaded into an extraction column for supercritical CO2 fluid extraction, and after dynamic extraction for 1 to 3 hours, separation treatment is performed to recover the polyphenol extract and the second solid; the polyphenol supernatant and the polyphenol extract are mixed and dried to obtain the avocado seed polyphenol; The supercritical treatment temperature is adjusted to 30-40° C. and the supercritical pressure is adjusted to 20-30 MPa so that the extraction fluid is in a supercritical state, and the flow rate of the entrainer is 0.2-0.6 mL / min.

2. The method for preparing avocado seed polyphenols according to claim 1, characterized in that: The extractant is one of 75% anhydrous ethanol, 85% anhydrous ethanol, 95% anhydrous ethanol, anhydrous ethanol, ethyl acetate, and acetone; The entrainer is one of 75% anhydrous ethanol, 85% anhydrous ethanol, 95% anhydrous ethanol, anhydrous ethanol, ethyl acetate and acetone.

3. The method for preparing avocado seed polyphenols according to claim 2, characterized in that: The extractant is 85% anhydrous ethanol, and the entrainer is 85% anhydrous ethanol; Alternatively, the extractant is ethyl acetate and the entrainer is ethyl acetate.

4. The method for preparing avocado seed polyphenols according to claim 1, characterized in that: In the pretreatment step, the outer skin of the avocado seeds is first removed and crushed into particles with a particle size of less than 0.2 cm; the particles are placed in an aerobic environment at 25 to 40° C. for 0 to 60 minutes for a spontaneous oxidation reaction; after the spontaneous oxidation reaction is completed, the particles are dried to a moisture content of between 10 and 15%, and crushed to pass through an 80-mesh sieve.

5. The method for preparing avocado seed polyphenols according to claim 1, characterized in that: In the first extraction step, the extractant is added to the particles at a mass ratio of 1:(15-20) between the particles and the extractant, and then the particles are stirred throughout the process. Ultrasonic treatment is performed for (2-5) minutes every (5-15) minutes, with an ultrasonic power of 400W-600W for 1-2 hours. The polyphenol supernatant and the first solid are filtered out, and the first solid is dried and then ultrafinely crushed to obtain avocado seed powder.

6. The method for preparing avocado seed polyphenols according to claim 1, characterized in that: In the second extraction step, the avocado seed powder is loaded into an extraction column, and then the extraction column is connected to the gas pipeline of a supercritical CO2 extractor, the supercritical treatment temperature is adjusted to 30-40°C and the supercritical treatment pressure is adjusted to 20-30MPa, so that the extraction fluid is in a supercritical state, the flow rate of the entrainer is adjusted to 0.2-0.6mL / min, and the dynamic extraction is performed for 1-3h; after the extraction is completed, the polyphenol extract and the second solid are recovered by vacuum separation; the polyphenol supernatant and the polyphenol extract are mixed, and freeze-dried to obtain avocado seed polyphenols.

7. A method for preparing avocado seed starch, characterized in that: The following steps are involved: Enzymatic starch production: the second solid is mixed with water, bromelain and papain are added, and stirred for a certain period of time; then the starch solid is separated to recover the starch solid, and the starch solid is washed and dried to obtain avocado seed starch; The second solid is prepared by the method for preparing avocado seed polyphenols according to any one of claims 1 to 6; the amount of bromelain and the amount of papain used are both 0.5% to 1% of the mass of the second solid.

8. The method for preparing avocado seed starch according to claim 7, characterized in that: In the enzymatic starch preparation step, the second solid is mixed with water in a mass ratio of 1: (10-15), and then bromelain and papain are added, and stirred for 1h-2h; then centrifugal separation is performed to recover the starch solid, and the starch solid is washed and dried to obtain avocado seed starch.

9. Avocado seed polyphenols, characterized in that: The avocado seed polyphenol is prepared by the preparation method of avocado seed polyphenol according to any one of claims 1 to 6.

10. Avocado seed starch, characterized in that: The avocado seed starch is prepared by the preparation method of avocado seed starch according to any one of claims 7 to 8.

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