CN105998109B - A kind of mango pit polyphenol extract and preparation method thereof - Google Patents

Abstract

The invention discloses a mango seed polyphenol extract and a preparation method thereof, belonging to the fields of food technology and biotechnology. The mango seed polyphenol extract is prepared by taking mango seeds as a raw material through alcohol extraction and a macroporous resin purification method. The mango seed polyphenol extract prepared by the invention has obvious bacteriostatic and antioxidant activity effects, and can be further researched and developed as an antioxidant health product or medicament.

Description

A kind of mango pit polyphenol extract and preparation method thereof

technical field

The invention relates to a mango pit polyphenol extract and a preparation method thereof, belonging to the fields of food technology and biotechnology.

Background technique

Mango (Latin name: Mangifera indica Linn), also known as citron fruit, stuffed fruit, honeydew, etc., belongs to the sumac family, mainly grows in tropical and subtropical coastal intertidal zones. The cultivation of mangoes is widely distributed. More than 70 countries in the world produce mangoes. The main provinces of my country are Guangxi, Guangdong, Taiwan, and Hainan, where mangoes are grown, and mangoes are rich in resources and varieties. In 2014, my country's mango planting area was 1.17 million hectares, with a total output of 880,000 tons. It is a large traditional tropical agricultural product in China. It has a huge potential for exporting foreign exchange and plays an important role in my country's foreign trade.

Mango is one of the famous tropical fruits, because of its delicate flesh (smooth and juicy), unique flavor (sweet taste), and rich in nutrients (rich in sugar, protein, crude fiber, vitamins, minerals and fats, etc.). Tropical fruit essence in one, known as the "king of tropical fruits". Mango has a very high medicinal value. Its peel can be used as medicine, which is a diuretic and dredging agent; nucleolus and mango leaves can also be used as medicine, which can detoxify, eliminate stagnation, and lower blood pressure. According to the analysis of traditional Chinese medicine, mango is a kind of fruit that is flat and sweet, quenches thirst and promotes fluid production, and has the effects of benefiting the stomach, relieving vomiting and preventing dizziness. Mangiferin contained in mango has obvious anti-peroxidation, protection of brain neurons and expectorant and cough.

As one of the few major provinces in my country that produces mangoes, Guangxi has the advantages of rich mango resources and a wide variety of varieties. However, at present, the market is mainly based on fresh sales from the origin, and a small amount is processed into juice, preserved fruit and pulp. , the mango deep processing industry is in urgent need of further development. Waste from mango production and processing used to be discarded or fed to livestock. In recent years, people have gradually carried out research on mango waste, including mango leaves, mango skin, mango stone, mango bark, etc., especially mango stone, which accounts for 20-60% of the total weight of mango, has attracted the attention of most researchers and entrepreneurs . According to reports, there are many kinds of chemical components in mango, mainly polyphenols, including mangiferin, quercetin, kaempferol, p-hydroxybenzoic acid, meta-coumaric acid, p-coumaric acid and ferulic acid, etc. Part of the active ingredient is located in the mango pit, which accounts for 20-60% of the total weight of the mango.

Mango core is rich in amino acids, polypeptides, proteins, phenols, flavonoids, polysaccharides, organic acids, saponins, tannins, flavonoids, anthraquinones, alkaloids, coumarins and lactones, triterpenes and steroids, alkaloids, A variety of active ingredients such as volatile oils and oils.

In recent years, more and more researches have been carried out in my country on active ingredients such as mango pit polyphenols, flavonoids, pyridone, terpenoids, and their antibacterial and antioxidant functions. However, there are still many problems. The separation effect is not obvious, the active ingredients obtained by identification are few, and the pharmacological effects of the active ingredients such as bacteriostatic and antioxidant effects are not clear.

The existing mango pit polyphenol extraction method also has problems such as low content of total polyphenols extracted and many organic solvent residues.

SUMMARY OF THE INVENTION

The purpose of the present invention is to make full use of natural resources, especially the characteristic fruit and vegetable resources of Guangxi, to provide a mango pit polyphenol extract and a preparation method thereof. The polyphenol extract has antibacterial and antioxidant properties and can be used for the preparation of health products or medicines . At the same time, in order to solve the problems of low total polyphenol content and more organic solvent residues in the existing mango pit polyphenol extraction methods, the method of the present invention prepares the mango pit polyphenol extract through alcohol extraction and macroporous resin purification method.

The preparation method of the mango pit polyphenol extract of the present invention comprises the following steps:

(1) dry and pulverize the mango pit;

(2) Extract the dried mango pit powder obtained in step (1) with an ethanol solution or methanol solution with an alcohol concentration of 20-90% (v/v) as a solvent, and then the obtained extract is concentrated under reduced pressure or vacuum-frozen dry to extract;

(3) dissolving the extract obtained in the step (2) with water, an ethanol solution with an alcohol concentration of less than 50% (v/v) or a methanol solution with an alcohol concentration of less than 50% (v/v) to remove the insoluble matter to obtain the above sample solution;

(4) Take a non-polar or weakly polar macroporous adsorption resin, load the sample solution obtained in step (3), first wash off impurities with water, and then use an eluent (ethanol solution or methanol solution) to elute , collect the eluate; the weight ratio of the macroporous adsorption resin and mango pit dry powder is 1:0.5-1:20;

(5) Concentrating the eluate collected in step (4) under reduced pressure or vacuum freeze-drying to obtain the mango pit polyphenol extract.

In an embodiment of the present invention, the extraction method in step (2) is to add 2-50L of ethanol solution or methanol solution with an alcohol concentration of 30%-80% per kilogram of mango core powder, and heat reflux or immersion or dipping extraction 2-5 times, 1-3h for each extraction during heat reflux extraction, 10-15 days for each extraction during immersion or dipping extraction, and the extracts are combined.

In one embodiment of the present invention, the extraction in step (2) is to extract mango pit powder with 10-95% (v/v, the same below) ethanol, preferably the concentration of ethanol is 20%-80% , most preferably 40%-60%.

In an embodiment of the present invention, the extraction in the step (2) is to extract the mango pit powder with ethanol for 1-4 times, most preferably 2-3 times.

In one embodiment of the present invention, in the extraction of the step (2), the ratio of solid to liquid (ratio of mango pit powder mass to ethanol solution, m/v) is 1:1-1:40, preferably 1:5 -1:30, more preferably 1:15-1:25.

In an embodiment of the present invention, in the extraction of the step (2), each extraction time is 0.5-4 h, preferably, the extraction time is 1-2 h.

In an embodiment of the present invention, the extraction in step (2) is performed at 40-60°C.

In an embodiment of the present invention, the specific process conditions of the step (2) are: 70% ethanol concentration, 1:25 material-to-liquid ratio, 60°C extraction temperature, and 120min extraction time.

In an embodiment of the present invention, the weight of the water, ethanol solution or methanol solution used for dissolving the extract in the step (3) is 3-50 times the weight of the extract.

In one embodiment of the present invention, the extraction method of step (3) is as follows: the extract obtained in step (2) is dissolved with 10 times the weight of the extract in water or 10% ethanol solution, and the insoluble matter is removed to obtain the sample solution.

In an embodiment of the present invention, the removal of insoluble matter in step (3) is performed by filtering or centrifuging.

In an embodiment of the present invention, the type of macroporous adsorption resin in step (4) is selected from HPD-722, HPD-826, LX-17, LX-26, LX-28, SP70, SP700, AB-8, DM21 or D101, preferably macroporous resin AB-8, DM21, D101, more preferably AB-8.

In an embodiment of the present invention, in step (4), washing with water first to remove impurities refers to eluting with 2-5BV of water to remove impurities.

In an embodiment of the present invention, the elution flow rate of the step (4) is an eluent flow rate of 1BV/h-10BV/h, preferably 2BV/h-5Bv/h.

In an embodiment of the present invention, in the step (4), the concentration of the ethanol solution or methanol solution of the eluent is 5%-95%, preferably 20%-80%; the elution volume of the eluent is 1-10 column volumes, preferably 3-8 column volumes.

In an embodiment of the present invention, the step (4) is to use an ethanol solution with an alcohol concentration of 40-70% for elution.

In one embodiment of the present invention, the purification method of step (4) is as follows: take the weakly polar macroporous resin with a weight ratio of mango stone powder of 1:5, and use the sample loading solution obtained in step (3) Load the sample, use 2-5BV of water to wash off impurities, and then use 3-10BV of 50% ethanol solution for elution, and collect the eluate obtained by elution with 50% ethanol solution.

In an embodiment of the present invention, in the step (4), the loading amount of the mango pit polyphenol extract (mass of mango pit powder: resin mass) is 0.5:1-20:1, preferably 1:1 -10:1.

In one embodiment of the present invention, the method is as follows: extracting mango pit polyphenols by heating and refluxing an ethanol solution, recovering the ethanol, concentrating, adsorbing the mango pit polyphenol extract on a macroporous resin, eluting, collecting and washing Deliquoring, concentrating and drying to obtain purified mango pit polyphenols.

In one embodiment of the present invention, the method is specifically: (1) dry and pulverize the mango pit; (2) weigh 1 kg of mango pit powder, add 20 L of ethanol solution with a concentration of 50%, and extract under reflux for three times. Reflux extraction for 1 h each time, combine the extracts, and concentrate the extracts under reduced pressure; (3) dissolve the extracts in water 10 times the weight of the extracts, filter out the insolubles, and obtain a sample loading solution; (4) put the sample The solution was loaded onto the column, the flow rate of the effluent was controlled to be 0.5BV/h, and the volume ratio of the sample loading to the resin AB-8 was 2:1, until all the extract entered the resin bed; 2-5BV of water was used for elution to remove impurities , then rinse the resin bed with 40-60% ethanol solution of 3-10BV, control the flow rate of the effluent to 2BV/h, and collect the eluent of the ethanol solution; (5) Concentrate the collected eluent to a certain volume under reduced pressure, carry out After spray drying, the purified product was obtained.

The present invention also claims the application of the mango pit polyphenol extract obtained according to the preparation method or purification method in the preparation of food, health care product or medicine.

The health care product or drug is a health care product or drug with antibacterial and antioxidant activity.

Beneficial effects of the present invention:

1. The present invention effectively improves the extraction efficiency of polyphenol substances by improving the extraction step of mango pit polyphenols, and the polyphenol content in the obtained extract can reach 18-20.9 mg/g (that is, 1 kg of dry powder of mango pit can contain 18-20.9 mg/g). The extraction yielded 18-20.9 g of polyphenols). Moreover, the method of the present invention uses water or ethanol as a solvent, which is cheap and non-toxic.

2. The present invention adopts the method of macroporous resin purification to further purify the extract. The purification method is easy and fast to operate, and the selected macroporous resin has the characteristics of large adsorption capacity and high desorption rate, and effectively removes impurities in the extract. The problem of organic solvent residue is prevented.

3. The mango pit polyphenol extract obtained according to the method of the present invention has a yield of 3.6-20.9 mg/g, and the obtained polyphenol extract has obvious scavenging effects on hydroxyl and superoxide radical anions.

Description of drawings

Fig. 1 is the standard curve diagram of gallic acid;

Fig. 2 is the scavenging effect of mango pit polyphenol extract on hydroxyl;

Fig. 3 is the scavenging effect of mango pit polyphenol extract on superoxide radical anion;

Figure 4 shows the scavenging effect of mango pit polyphenol extract on DPPH.

Detailed ways

The technical content of the present invention will be further described below in conjunction with the embodiments. The following embodiments are illustrative, not restrictive, and the protection scope of the present invention cannot be limited by the following embodiments.

The equipment, chemical reagents and detection methods used in each embodiment are as follows:

Instruments and equipment: FW100 high-speed universal pulverizer, Tianjin Test Instrument Co., Ltd.; UPC-Ⅱ-20T Youpu Series Ultrapure Water Device, Sichuan Youpu Ultrapure Technology Co., Ltd.; SQP Electronic Balance, Sartorius Scientific Instruments (Beijing) Co., Ltd.; ZWYR-D2403 Oscillating Incubator, Shanghai Zhicheng Analytical Instrument Manufacturing Co., Ltd.; SHZ-DⅢ Yuhua Brand Circulating Water Vacuum Pump, Gongyi Yuhua Instrument Co., Ltd.; HHS-6S Electronic Constant Temperature Stainless Steel Water Bath Pot, Shanghai Yichang Instrument Yarn Sieve Factory; UV1901PC UV-Vis Spectrophotometer, Shanghai Aoyan Scientific Instrument Co., Ltd.;

Chemical reagents: anhydrous sodium carbonate, methanol, ethanol, acetone, ethyl acetate, Folin phenol reagent, gallic acid standard, various types of macroporous resins, etc. are all provided by Liuzhou Suli Co., Ltd.

Assay:

Establishment of gallic acid standard curve: Accurately weigh 50 mg of gallic acid, dilute to 50 ml with 70% ethanol solution, and obtain a standard solution of gallic acid with a concentration of 1.0 mg/ml. Take 9 50ml volumetric flasks, pipette 0, 1, 2, 3, 4, 5, 7.5, 10, 12.5ml of the above gallic acid standard solution respectively, dilute them to make 0, 0.02, 0.04, 0.06, 0.08, Standard solutions of 0.10, 0.15, 0.20, 0.25 mg/ml. Absorbance values were determined by the Folin method. Taking the gallic acid concentration as the abscissa and the absorbance value at 765nm as the ordinate, draw a standard curve and perform regression analysis. The standard curve for the determination of total phenols by the Folin-phenol method (Figure 1). The results show that gallic acid has a good linear relationship with its absorbance value in the concentration range of 0-0.25mg/ml. The linear regression equation in this range is: y=4.2974x+0.056, R ² =0.9994.

Determination of total phenolic content: Accurately pipette 2.0ml of the solution to be tested into a 50ml volumetric flask, add 1.0ml of Folin reagent, fully mix it and let it stand for 2min, add 10ml of 7% sodium carbonate solution, fully mix and dilute to volume with distilled water , and placed in a constant temperature water bath at 30 °C for 1 h with constant shaking. Take out, and measure the absorbance value at a wavelength of 765 nm (with distilled water as a blank control). Calculate the polyphenol content in the liquid to be tested according to the standard curve and its regression equation, and then calculate the content of polyphenols in the powder sample of 1 g mango pit after drying and crushing (calculated as gallic acid, mg/g, that is, polyphenols Content mg/g of mango pit powder).

Determination of Antioxidant Activity:

Determination of the ability of scavenging hydroxyl radicals: Take the sample solutions of different mass concentrations in 5 test tubes, add ferrous sulfate and hydrogen peroxide to each, let stand, add salicylic acid after 10 minutes, and measure the absorbance A at 510 nm after standing _1. Use distilled water instead of salicylic acid to measure the absorbance value A ₂ according to the above method, use distilled water to replace the liquid to be tested and measure the absorbance value A ₀ according to the above method, V _C is used as a control, and the scavenging of hydroxyl radicals by polyphenols is calculated according to the following formula Rate: hydroxyl radical (%)=[A ₀ -(A ₁ -A ₂ )]/(A ₀ )*100(%).

Determination of superoxide anion free radical scavenging ability: take 4.5ml Tris-HCl buffer solution, preheat in 20℃ water bath for 20min, add 1ml sample solution of different concentrations and 0.4ml 25mmol/L pyrogallol solution respectively, mix well Then, react in a 25°C water bath for 5 min, then add 1 ml of 8 mol/L HCl solution to terminate the reaction, and then measure the absorbance A ₁ at 325 nm, replace the sample solution with distilled water for the blank, and measure the absorbance A ₀ . The scavenging rate of samples to superoxide anion radicals: scavenging rate (%)=(1-A ₁ /A ₀ )*100 (%).

Determination of DPPH scavenging ability: Take 2.0 mL of different mass concentrations (1.0, 5.0, 10.0, 15.0, mango pit active ingredient sample solution respectively in 5 test tubes, add 2 mL of DPPH (0.04 mg/ml) to each, and let stand for 20 min and detect at 517 nm Absorbance A ₁ ; replace DPPH with absolute ethanol, measure absorbance A ₂ according to the above-mentioned method; do blank group measure absorbance A ₀ by the above-mentioned method, take _VC as a control, the scavenging rate of sample to DPPH free radical: DPPH scavenging rate (% )=[1-(A ₁ -A ₂ )/A ₀ ]*100(%).

Embodiment 1: the preparation method of the mango pit polyphenol extract of the present invention

(1) Extraction of polyphenols from mango pits: dry and pulverize mango pits, weigh 1 kg of mango pit powder, add 20 L of ethanol with a concentration of 50%, extract three times under heat reflux, extract under heat reflux for 1 hour each time, combine the extracts, and concentrate the extracts under reduced pressure Juice extract, the extract is dissolved in water 10 times the weight of the extract, and the insolubles are filtered out to obtain a sample loading solution with a total polyphenol content of 18-20.9g/kg mango core powder.

(2) Column loading: the mango pit polyphenol extract was loaded onto the column, the flow rate of the effluent was controlled to be 0.5BV/h, and the volume ratio of the loading volume to the resin AB-8 was 2:1, until all the extract entered the resin bed.

(3) Elution: use 2-5BV of water to elute to remove impurities, then use 3-10BV of 5%-80% ethanol to rinse the resin bed, control the flow rate of the effluent to 2BV/h, and collect the eluent, Respectively concentrated under reduced pressure to a certain volume, and spray-dried to obtain purified products of different components, wherein the total polyphenol content of all components is about 20.9g/kg, and in the purified product, the polyphenol content in the water eluent is about 2.2g/kg, the polyphenol content in the 5% ethanol eluent is about 3.8g/kg, the polyphenol content in the 10% ethanol eluent is about 4.8g/kg, and the polyphenol content in the 20% ethanol eluent is about It is 8.04g/kg, the polyphenol content in 30% ethanol eluent is about 8.5g/kg, the polyphenol content in 40% ethanol eluent is about 13.3g/kg, and the polyphenol content in 50% ethanol eluent is about 13.3g/kg About 15.2g/kg, the content of polyphenols in 60% ethanol eluent is about 13.7g/kg, the content of polyphenols in 70% ethanol eluent is about 11.1g/kg, and the polyphenol content in 80% ethanol eluent is about 11.1g/kg The content is about 9.9g/kg.

Example 2: Select different conditions for extraction

Using the method provided by the present invention, the extraction experiment of mango pit polyphenols (sample is 1 g of mango pit dried and pulverized) is carried out under different conditions. The L ₉ (3 ⁴ ) orthogonal test (Table 1) was performed on the factors, and the orthogonal result analysis was shown in Table 2.

Table 1 Design of the orthogonal test table for the extraction process of polyphenols from mango core

Table 2 Orthogonal experimental results of mango pit polyphenol extraction process L ₉ (3 ⁴ )

Table 3 Mango kernel polyphenol extraction verification test

It can be seen from Table 2 that the order of the influence of various factors on the extraction of mango pit polyphenols is: B>A>D>C, that is, the ratio of solid to liquid>ethanol concentration>extraction time>extraction temperature, in which the ratio of solid to liquid affects the extraction of polyphenols from mango pit The effect was the most significant, followed by ethanol concentration and extraction time. The effect was the same, and the extraction temperature had the least significant effect on the extraction of polyphenols from mango pits. From the orthogonal test results, it can be concluded that the best test combination is A ₃ B ₃ C ₃ D ₃ , that is, the concentration of ethanol is 70%, the ratio of solid to liquid is 1:25, the extraction temperature is 60°C, and the extraction time is 120min. It is not in the combination of orthogonal tables, so a verification test (Table 3) is required. From the verification test results, it can be seen that the extraction content of polyphenols is 5.36 mg/g, so the optimal process conditions for the extraction of polyphenols from mango kernels is ethanol concentration 70 %, the ratio of solid to liquid is 1:25, the extraction temperature is 60 °C, and the extraction time is 120 min.

Example 3: Macroporous resin purification process

Select different resin types, sample concentration, adsorption rate, sample loading, ethanol elution concentration, elution rate and other factors to carry out single-factor experiments respectively to determine the optimal value of each factor.

Preparation of sample solution: Weigh 20 g of mango pit powder, according to ethanol concentration of 70%, solid-liquid ratio of 1:25, extraction temperature of 60 °C, each extraction for 1 hour, and three times of extraction. After extraction, the filtrates were combined and concentrated to dryness under reduced pressure. Vacuum dry to constant weight, accurately weigh an appropriate amount of dry paste, dissolve it in a 10ml volumetric flask with methanol or ethanol solution, add methanol or ethanol solution to dilute to the mark, and shake well for later use. Pipette 1ml of filtrate into a 10ml volumetric flask, add methanol or ethanol solution to dilute to the mark, and shake well. The polyphenol content was determined as described above.

Static adsorption: Take 10 g each of 10 pretreated macroporous adsorption resins, add 10 ml of mango pit polyphenol solution to each, shake once every 10 minutes, and take the resin-adsorbed solution after 2 hours to measure the polyphenol content, and calculate each Adsorption rate of mango pit polyphenols by resin.

Static desorption:

Filter and drain the statically adsorbed resin, add 30 ml of 50% ethanol for desorption, shake once every 10 min, and after 2 h, take the resin-adsorbed solution to measure the polyphenol content, and calculate the desorption rate of mango core polyphenols by each resin. See Table 4. It can be seen from the table that the adsorption rates of the 10 resins are all large (greater than 90%), and the desorption rates are all greater than 40%. Among them, the desorption rates of AB-8, DM21 and D101 are all greater than 60%. DM21, D101 do dynamic adsorption experiments.

Table 4 Adsorption and desorption rates of mango pit polyphenols by macroporous resin

Dynamic adsorption:

Take 15mL of each of the three preferred resins AB-8, DM21, and D101 for static adsorption in the column, add mango core polyphenol extract to the top of the column, carry out dynamic adsorption at a flow rate of 0.5BV/h, and collect according to the volume of the resin. The effluent was measured for polyphenol content, and the adsorption rate of mango pit polyphenols by each resin was calculated. The results are shown in Table 5, indicating that when AB-8 and D101 reach saturation adsorption, the total amount of adsorbed polyphenols is larger than that of DM21, which is selected for desorption experiments.

Table 5 Dynamic adsorption results of mango pit polyphenols by macroporous resin

Dynamic desorption: take 20ml of each of the treated AB-8 and D101 resins in the column, add mango core polyphenol extract (10.5mg/g) to the top of the column respectively, carry out adsorption at a flow rate of 0.5BV/h, use 60 % ethanol was eluted at a flow rate of 2BV/h, and the eluate was collected according to the volume of the resin, the polyphenol content was determined, and the adsorption rate of mango core polyphenols by each resin was calculated. The results are shown in Table 6. It can be seen that AB-8 has better desorption effect than D101, and it is selected as the best resin.

Table 6 Dynamic desorption results of mango core polyphenols by macroporous resin

Ethanol concentration: take 10 parts of 20ml of the treated AB-8 resin in the column, add mango kernel polyphenol extract on the top of the column, carry out adsorption at a flow rate of 0.5BV/h, and then use 5BV of water, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80% ethanol was eluted separately at a flow rate of 2BV/h, and the eluate was collected according to the volume of the resin, and the polyphenol content was determined. The results are shown in Table 7, it can be seen that when the ethanol elution concentration is 40%-70%, the eluted total polyphenol yield is the highest.

Table 7 Investigation of ethanol elution concentration

Example 4: Preparation method of mango pit polyphenol extract

Dry and pulverize the mango pits, weigh 1 kg of mango pit powder, add 20 L of ethanol with a concentration of 50%, extract three times under heat reflux, and extract under heat reflux for 1 h each time, combine the extracts, concentrate the extracts under reduced pressure, and extract the extract. 10 times the weight of the paste was dissolved in water, and the insolubles were filtered out to obtain a sample loading solution with a total polyphenol content of 18-20.9 g/kg.

The mango pit polyphenol extract was loaded onto the column, the flow rate of the effluent was controlled to be 0.5BV/h, and the volume ratio of the loading volume to the resin AB-8 was 2:1, until all the extract entered the resin bed. Use 2-5BV of water to remove impurities, then use 3-10BV of 50% ethanol to rinse the resin bed, control the flow rate of the effluent to 2BV/h, collect the eluent, concentrate to a certain volume under reduced pressure, and spray dry to obtain purification. The product has a polyphenol content of 14-15.2 g/kg.

Example 5: Preparation method of mango pit polyphenol extract

Dry and pulverize the mango pits, weigh 1 kg of mango pit powder, add 20 L of ethanol with a concentration of 50%, extract three times under heat reflux, and extract under heat reflux for 1 h each time, combine the extracts, concentrate the extracts under reduced pressure, and extract the extract. 10 times the weight of the paste was dissolved in water, and the insolubles were filtered out to obtain a sample loading solution with a total polyphenol content of 18-20.9 g/kg.

The mango pit polyphenol extract was loaded onto the column, the flow rate of the effluent was controlled to be 0.5BV/h, and the volume ratio of the loading volume to the resin AB-8 was 2:1, until all the extract entered the resin bed. Use 2-5BV of water to remove impurities, then use 3-10BV of 40% ethanol to rinse the resin bed, control the flow rate of the effluent to 2BV/h, collect the eluent, concentrate under reduced pressure to a certain volume, and spray dry to obtain purification. The product has a polyphenol content of 12-13.3 g/kg.

Example 6: Preparation method of mango pit polyphenol extract

Dry and pulverize the mango pits, weigh 1 kg of mango pit powder, add 20 L of ethanol with a concentration of 50%, extract three times under heat reflux, and extract under heat reflux for 1 h each time, combine the extracts, concentrate the extracts under reduced pressure, and extract the extract. 10 times the weight of the paste was dissolved in water, and the insolubles were filtered out to obtain a sample loading solution with a total polyphenol content of 18-20.9 g/kg.

The mango pit polyphenol extract was loaded onto the column, the flow rate of the effluent was controlled to be 0.5BV/h, and the volume ratio of the loading volume to the resin AB-8 was 2:1, until all the extract entered the resin bed. Use 2-5BV of water to remove impurities, then use 3-10BV of 70% ethanol to rinse the resin bed, control the flow rate of the effluent to 2BV/h, collect the eluent, concentrate under reduced pressure to a certain volume, and spray dry to obtain purification. The product, the polyphenol content is 10-11.1 g/kg.

Example 7: Antibacterial and antioxidant properties of mango pit polyphenol extract obtained by the method of the present invention

Determination of antibacterial effect:

的培养皿中，每皿15-20mL培养基，待平板冷却后，每皿中加入0.5mL菌悬液，用三角玻璃涂棒均匀涂成薄板备用。将无菌滤纸片浸入配好的药液其中12h，沥干。每一含菌平板上，呈正三角形排布3片带药无菌纸片，细菌于37℃条件下培养24h，啤酒酵母和黑曲霉于30℃条件下培养5d，测其抑菌圈直径。根据抑菌圈大小来确定其抑菌效果。以不加芒果核多酚溶液只加菌悬液为对照。结果显示，本发明得到的芒果核多酚提取物具有强烈的抑菌效果。The test strain was activated on the slant medium, and 1-2 rings were picked with an inoculation ring and shaken in a 50mL sterile saline flask for 10min (with several glass beads in it), and the bacterial suspension was required to be about 10 ⁸ CFU/mL Spare left and right. Weigh a certain amount of the mango pit polyphenol extract samples obtained in Examples 4-6 which were vacuum freeze-dried, and dissolve the samples with sterile 40% ethanol aqueous solution on the ultra-clean workbench to the required concentration, for later use. The freshly prepared medium is sterilized at 121°C. When the medium is cooled to 50-60°C, pour the medium into the sterilized medium on the ultra-clean workbench.

Add 0.5 mL of bacterial suspension to each dish after the plate is cooled, and evenly coat it into a thin plate with a triangular glass coating rod for later use. Immerse the sterile filter paper in the prepared medicinal solution for 12h and drain. On each bacteria-containing plate, three sterile paper sheets with medicine were arranged in a regular triangle. The bacteria were cultured at 37°C for 24 hours, and the yeast and Aspergillus niger were cultured at 30°C for 5 days. The diameter of the inhibition zone was measured. The bacteriostatic effect was determined according to the size of the inhibition zone. To control without adding mango pit polyphenol solution and only adding bacterial suspension. The results show that the mango pit polyphenol extract obtained by the present invention has a strong antibacterial effect.

Antioxidant activity of mango pit polyphenols:

Hydroxyl is the most active active oxidative free radical, which can induce oxidative damage in the body, and its scavenging rate is often an important indicator of the antioxidant effect of the reaction drug. Similarly, superoxide anion is also the main reactive oxygen radical in the organism, and the lipid peroxidation caused by it is an important cause of aging, cardiovascular disease and tumorigenesis. The scavenging capacity of mango pit polyphenols hydroxyl radicals is shown in Figure 2. The scavenging capacity increases with the increase of polyphenol concentration. When the concentration of mango pit polyphenols reaches 0.6 mg/ml, the scavenging rate can reach about 90%, indicating that mango pits Polyphenols have a strong scavenging effect on hydroxyl radicals. The superoxide anion free radical scavenging ability of mango pit polyphenol is shown in Figure 3. The higher the concentration of mango pit polyphenol, the stronger the superoxide anion free radical scavenging ability. When the concentration of mango pit polyphenol reaches 0.5-0.6mg/ mL, the clearance capacity tends to be stable, and the clearance rate reaches 83%, but the clearance capacity is weaker than that of Vc. The results of DPPH free radical scavenging ability of mango kernel polyphenol extract are shown in Figure 4. The scavenging ability increases with the increase of polyphenol concentration. When the concentration reaches 0.6 mg/mL or more, the scavenging rate of DPPH free radical can reach 90%. %, indicating that mango pit polyphenols have a strong scavenging effect on DPPH free radicals. The above data show that mango pit polyphenols have good scavenging ability to hydroxyl radicals and superoxide anion free radicals. The research results will provide reference for the comprehensive utilization of mango pit resources and the development of corresponding functional foods.

The content of the present invention has been described in detail above through the embodiments, but the content is only a preferred embodiment of the present invention and cannot be considered to limit the scope of implementation of the present invention. Anyone who is familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention should be defined by the claims.

Claims (5)

1. a preparation method of mango pit polyphenol extract, is characterized in that, described method comprises the following steps: (1) Dry and crush the mango pit; (2) Using an ethanol solution with an alcohol concentration of 70% v/v as a solvent, extract the dried mango pit powder obtained in step (1). Time 120min; then the obtained extract is concentrated under reduced pressure or vacuum freeze-dried to extract; (3) dissolving the extract obtained in step (2) with water, an ethanol solution with an alcohol concentration of less than 50% v/v, or a methanol solution with an alcohol concentration of less than 50% v/v, to remove insoluble matter to obtain a sample loading solution; (4) Take the weakly polar macroporous adsorption resin with a weight ratio of 1:5 to mango stone powder, load the sample solution obtained in step (3), first wash away impurities with 2-5 BV water, and then use 3 50% eluent of -10 BV is eluted, and the eluent obtained by elution of 50% ethanol solution is collected; (5) Concentrating the eluate collected in step (4) under reduced pressure or vacuum freeze-drying to obtain the mango pit polyphenol extract. 2 . The method according to claim 1 , wherein the macroporous adsorption resin in the step (4) is AB-8, DM21 or D101. 3 . 3. The method according to claim 1, characterized in that, in the step (2), the extraction method is to add 2-50L ethanol solution with an alcohol concentration of 70% per kilogram of mango pit powder, and heat reflux or dipping or dipping. Extract 2-5 times, extract 1-3 hours each time during heat reflux extraction, and extract each time for 10-15 days during immersion or dipping extraction, and combine the extracts. 4 . The method according to claim 1 , wherein the extraction in the step (2) is carried out at 60° C., the extraction times are 1-4 times, the extraction time is 0.5-4h each time, and the quality of mango stone powder is 1-4. 5 . The ratio to the volume of ethanol solution is 1:25. 5. The mango pit polyphenol extract obtained by the method according to claim 1.

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