Olive leaf water extract and preparation method thereof

Abstract

The present invention provides an olive leaf water extract and a preparation method thereof. The present invention provides an olive leaf water extract preparation method, comprising: softening the olive leaves and then removing bitterness to obtain de-astringent olive leaves; crushing the de-astringent olive leaves and then enzymolyzing them with a composite enzyme to obtain an olive leaf hydrolysate; the composite enzyme includes papain, pectinase, cellulase and neutral protease; and ultrasonically treating the olive leaf hydrolysate to obtain an olive leaf water extract. The preparation method of the present invention can fully extract the active ingredients in the olive leaves, so that the active ingredients in the olive leaves are fully absorbed, thereby achieving the effect of enhancing physical fitness.

Description

Technical Field

[0001] The invention specifically relates to an olive leaf water extract and a preparation method thereof. Background Art

[0002] Olives are cold-pressed into olive oil, which has high nutritional value and market prospects, but olive leaves are not fully utilized. Some are used to produce feed, but most are burned and composted, causing environmental pollution and waste of resources. Studies have shown that olive leaves are rich in nutrients, such as olive polyphenols, flavonoids, oleuropein, dietary fiber, reducing sugars, hydroxytyrosol, lignans, flavonoids, etc. The content of some of these substances in olive leaves is even higher than that in olives and olive oil. Most of these substances have multiple pharmacological activities such as anti-inflammatory and antibacterial, hypoglycemic, hypolipidemic, antioxidant and antiviral. There are also data showing that they have an inhibitory effect on the growth of three foodborne pathogens, Listeria monocytogenes, Escherichia coli and Salmonella enterica, and can provide a beneficial intestinal microecological environment.

[0003] Therefore, how to provide a preparation method for extracting the active ingredients of olive leaves is of great significance to the full utilization of olive leaves. Summary of the invention

[0004] In view of the defects in the prior art, the object of the present invention is to provide a method for preparing an olive leaf water extract, which can make rational use of the active ingredients in the olive leaf.

[0005] The objective of the present invention is achieved through the following technical solutions:

[0006] The present invention provides a method for preparing an olive leaf water extract, comprising the following steps:

[0007] The olive leaves are softened and then debittered to obtain de-astringent olive leaves;

[0008] The de-astringent olive leaves are crushed and then enzymolyzed with a composite enzyme to obtain an olive leaf hydrolyzate; the composite enzyme comprises papain, pectinase, cellulase and neutral protease;

[0009] The olive leaf enzymatic hydrolysate is subjected to ultrasonic treatment to obtain an olive leaf water extract.

[0010] Preferably, the mass ratio of papain, pectinase, cellulase and neutral protease in the composite enzyme is 1:1:2:1;

[0011] The enzymatic activity of the papain is 2400U / mg; the enzymatic activity of the pectinase is 500U / g; the enzymatic activity of the cellulase is 10000U / g; the enzymatic activity of the neutral protease is 70U / mg;

[0012] The mass ratio of the olive leaf to the complex enzyme is (10-50): (0.5-1.5);

[0013] The temperature of the enzymolysis is 40° C. to 70° C.; the time of the enzymolysis is 2 to 4 hours.

[0014] Preferably, the softening comprises soaking the olive leaves in an aqueous solution of a green preserving agent; the soaking time is 0.5 to 2 hours;

[0015] The greening agent comprises ascorbic acid, citric acid and sodium bicarbonate; the mass ratio of ascorbic acid, citric acid and sodium bicarbonate in the greening agent is (3-9):(3-5):1;

[0016] In the greening agent aqueous solution, the mass ratio of the greening agent to water is (5-50):(500-4000);

[0017] The debittering agent comprises a brine solution containing lime; the mass concentration of NaCl in the brine solution containing lime is 4% to 8%; the mass concentration of lime in the brine solution containing lime is 0.04% to 0.06%;

[0018] The debittering time is 3 to 4 hours.

[0019] Preferably, the power of the ultrasound is 600-1000W, the time of the ultrasound is 15-45min, and the temperature of the ultrasound is 50°C-70°C.

[0020] The present invention provides an olive leaf water extract prepared by the preparation method described in the above technical solution.

[0021] Beneficial effects of the present invention:

[0022] The present invention provides a method for preparing an olive leaf water extract, comprising the following steps: softening the olive leaves and then removing the bitterness to obtain astringent olive leaves; crushing the astringent olive leaves and then performing enzymolysis with a composite enzyme to obtain an olive leaf hydrolysate; the composite enzyme includes papain, pectinase, cellulase and neutral protease; and ultrasonically treating the olive leaf hydrolysate to obtain an olive leaf water extract. The method for preparing the olive leaf water extract provided by the present invention can fully extract the active ingredients in the olive leaves, so that they can be fully absorbed in the body and enhance physical fitness. The olive leaf water extract prepared by the preparation method of the present invention is rich in olive polyphenols, flavonoid compounds, oleuropein, dietary fiber, reducing sugar, hydroxytyrosol, lignans and flavonoids, etc., and the olive leaf water extract has multiple pharmacological activities such as anti-inflammatory and antibacterial, hypoglycemic, hypolipidemic, anti-oxidative and antiviral. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative labor.

[0024] Figure 1 This is a graph showing the free radical scavenging rate test results of olive leaf extracts obtained by enzymatic extraction, conventional water extraction and alcohol extraction;

[0025] Figure 2 This is a photograph of the olive leaves in Example 1-2 before mixing with pure water for juicing;

[0026] Figure 3 This is a photographic result of the olive leaves of Comparative Example 1-9 before being mixed with purified water for juicing;

[0027] Figure 4 A graph showing the chlorophyll content in the olive leaf water extracts prepared in Example 1-1, Example 1-2 and Comparative Examples 1-6 to 1-8;

[0028] Figure 5 The results of active ingredient detection of olive leaf water extract prepared in Examples 1-2 to 1-3 and Comparative Examples 1-4 and 1-5;

[0029] Figure 6 This is the morphology of the olive leaf water extract prepared in Example 1-2. DETAILED DESCRIPTION

[0030] The present invention provides a method for preparing an olive leaf water extract, comprising the following steps:

[0031] The olive leaves are softened and then debittered to obtain de-astringent olive leaves;

[0032] The de-astringent olive leaves are crushed and then enzymolyzed with a composite enzyme to obtain an olive leaf hydrolyzate; the composite enzyme comprises papain, pectinase, cellulase and neutral protease;

[0033] The olive leaf enzymatic hydrolysate is subjected to ultrasonic treatment to obtain an olive leaf water extract.

[0034] The method comprises the following steps: softening the olive leaves and then removing the bitterness to obtain the de-astringent olive leaves.

[0035] In the present invention, the olive leaves are preferably fresh, tender green olive leaves, more preferably fresh tender green leaves picked between February and May. In the present invention, the olive leaves are preferably olive leaves, more preferably olive leaves planted in Longnan, Gansu. After obtaining the olive leaves, the present invention preferably softens the olive leaves after washing. The present invention does not specifically limit the washing method, and conventional washing methods in the art can be used. The washing described in the present invention is preferably rinsed with clean water for multiple times. The softening described in the present invention is preferably carried out in an aqueous solution of a greening agent. In the present invention, the greening agent preferably includes ascorbic acid, citric acid and sodium bicarbonate; the mass ratio of ascorbic acid, citric acid and sodium bicarbonate in the greening agent is preferably (3-9):(3-5):1, further preferably (4-8):(3-5):1, and more preferably 6:4:1. In the present invention, the mass ratio of the greening agent to water in the greening agent aqueous solution is preferably (5-50): (500-4000), more preferably (7-45): (800-3000), and more preferably 11:1800. The soaking time of the present invention is preferably 0.5-2h, more preferably 0.5-1.5h, and more preferably 1h. The addition of the greening agent during softening can achieve the purpose of not destroying the overall structure of the cell wall; in addition, during the heat treatment, chlorophyll is easily destroyed, causing the olive leaves to lose their bright green color and form brown. The addition of the greening agent here can ensure the stability of the chlorophyll.

[0036] After softening, the present invention debitters the softened olive leaves. The debittering agent of the present invention preferably includes a lime-containing aqueous solution, more preferably a lime-containing brine solution. In the present invention, the mass fraction of NaCl in the lime-containing brine solution is preferably 4% to 8%, more preferably 6%. In the present invention, the mass concentration of lime in the lime-containing brine solution is preferably 0.04% to 0.06%. , More preferably 0.05%. In the present invention, the lime is preferably food-grade lime powder, and the present invention has no special restrictions on the source of lime, and commercial products can be used. The food-grade lime powder in the present invention is purchased from Taobao.com. The debittering of the present invention preferably immerses the olive leaves in a lime brine solution for immersion. The debittering time of the present invention is preferably 3 to 4 hours, more preferably 3.5 hours. After debittering is completed, the present invention preferably washes and dries the debittered olive leaves to obtain de-astringent olive leaves. The present invention has no special restrictions on the washing and drying methods, and conventional washing and drying methods in the art can be used. The washing of the present invention is preferably washed with pure water.

[0037] After obtaining the de-astringent olive leaves, the present invention crushes the de-astringent olive leaves and then uses a composite enzyme for enzymolysis to obtain an olive leaf hydrolysate. Before enzymolysis, the present invention preferably mixes the de-astringent olive leaves with water and then crushes them to obtain olive leaf juice. In the present invention, the mass ratio of the olive leaves to water is preferably 1: (8-10), more preferably 1: 9. In the present invention, the water is preferably purified water. The crushing of the present invention preferably squeezes the olive leaves into juice. The crushing of the present invention is preferably carried out using a wall breaking machine. After obtaining the olive leaf juice, the present invention uses a composite enzyme for enzymolysis of the olive leaf juice. In the present invention, the composite enzyme preferably includes papain, pectinase, cellulase and neutral protease. In the present invention, the mass ratio of papain, pectinase, cellulase and neutral protease in the composite enzyme is preferably 1: 1: 2: 1. In the present invention, the enzymatic activity of the papain is preferably 2400U / mg; the enzymatic activity of the pectinase is preferably 500U / g; the enzymatic activity of the cellulase is preferably 10000U / g; the enzymatic activity of the neutral protease is preferably 70U / mg. In the present invention, the neutral protease preferably includes subtilisin and serrapeptase. In the present invention, the papain, pectinase, cellulase and neutral protease are preferably purchased from Nanning Dongheng Huadao Biotechnology Co., Ltd. In the present invention, the mass ratio of the olive leaf and the compound enzyme is preferably (10-50): (0.5-1.5), more preferably (10-30): (0.5-1.5), and more preferably 20:1. The temperature of the enzymolysis of the present invention is preferably 40℃~70℃, more preferably 50℃~70℃, and more preferably 60℃; the time of the enzymolysis is preferably 2-4h, and more preferably 3h. The present invention uses a composite enzyme to hydrolyze olive leaves, which can quickly and efficiently hydrolyze the protein in the olive leaves into amino acids and polypeptides, increase the content of active ingredients, and also make the body absorb better. In the present invention, the pectinase has a good effect on decomposing pectin; papain is a pure natural endopeptidase extracted by modern bioengineering technology, contains sulfhydryl (-SH), catalyzes protein hydrolysis, has the dual activity of protease and esterase, and has a strong hydrolysis ability for animal and plant proteins, polypeptides, esters and amides; subtilisin and serrapeptidase in the neutral protease are biological enzyme preparations refined by biotechnology after culturing and fermenting Bacillus subtilis, and can hydrolyze macromolecular proteins into products such as polypeptides and amino acids; the cellulase mainly destroys the cell wall structure of olive leaves by enzymatic hydrolysis, thereby hydrolyzing cellulose in the cell wall of olive leaves and reducing the mass transfer resistance during the extraction process. The present invention selects composite enzymes in the preparation process of olive leaf extract mainly because olive leaves contain dietary fiber, reducing sugar, oleuropein, etc., and have a complex structure, while the enzymatic hydrolysis effect of monomeric enzymes is single. Therefore, composite enzymes are selected to catalyze the decomposition of macromolecules.

[0038] After obtaining the olive leaf hydrolysate, the present invention preferably subjects the olive leaf hydrolysate to ultrasonic treatment to obtain an olive leaf water extract.

[0039] The power of the ultrasound of the present invention is preferably 600-1000W, more preferably 800W; the time of the ultrasound is preferably 15-45min, more preferably 20-40min, more preferably 30min; the temperature of the ultrasound is preferably 40°C-70°C, more preferably 40°C-60°C, more preferably 50°C. After the ultrasound is completed, the present invention preferably also includes filtering, taking the supernatant to obtain the olive leaf water extract. The present invention does not specifically limit the filtering method, and the conventional filtering method in the art can be used. The ultrasound of the present invention is preferably carried out in a microwave ultrasound instrument. The present invention adopts ultrasound-assisted extraction technology, and utilizes the cavitation and dispersion of the ultrasonic pulser in the sample to directly and indirectly act on the solvent and solute, which is beneficial to the mass transfer of the solvent and promotes the release of bioactive substances in olive leaves.

[0040] The olive leaf water extract prepared by the preparation method of the technical scheme is rich in olive polyphenols, flavonoids, oleuropein, dietary fiber, reducing sugar, hydroxytyrosol, lignans and flavonoids, etc., some of which are even higher in content in olive leaves than in olive fruit and olive oil, and most of these substances have multiple pharmacological activities such as anti-inflammatory and antibacterial, hypoglycemic and blood lipid, anti-oxidation, and antiviral. The olive leaf water extract of the present invention has an inhibitory effect on the growth of three foodborne pathogens of Listeria monocytogenes, Escherichia coli and Salmonella enterica, thereby providing a beneficial intestinal microecological environment. The preparation method of the olive leaf water extract provided by the present invention can significantly improve the free radical scavenging rate of the olive leaf extract. The preparation method of the olive leaf water extract provided by the present invention can better protect the chlorophyll in the olive leaf and significantly increase the content of chlorophyll in the product. The preparation method of the olive leaf water extract provided by the present invention can significantly increase the content of active ingredients such as oleuropein, hydroxytyrosol, verbascoside, luteolin and gallic acid in the olive leaf.

[0041] The present invention provides an olive leaf water extract prepared by the preparation method described in the above technical solution. In the present invention, the olive leaf water extract includes oleuropein, hydroxytyrosol, verbascoside, luteolin and gallic acid. In the present invention, the olive leaf water extract contains a high content of chlorophyll and has a high free radical scavenging rate.

[0042] In order to further illustrate the present invention, the technical solution provided by the present invention is described in detail below in conjunction with the accompanying drawings and embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0043] Example 1-1

[0044] A method for preparing an olive leaf water extract comprises the following steps:

[0045] The olive leaves come from olive trees planted in Longnan, Gansu Province. Fresh green leaves picked in February were selected. 10g of fresh green olive leaves were rinsed with clean water several times, and then soaked in pure water with greening agent added for 1 hour to soften the olive leaves.

[0046] The greening agent is greening agent component 1, and the mass ratio of ascorbic acid, citric acid and sodium bicarbonate in the greening agent component 1 is 3:3:1. The preparation method of the greening agent pure water is: add 0.3g of ascorbic acid, 0.3g of citric acid and 0.1g of sodium bicarbonate into 80mL of pure water.

[0047] The olive leaves soaked in the above step are immersed in salt water with lime for 3 hours to remove bitterness and astringency, and then washed with purified water and dried to obtain de-astringent olive leaves;

[0048] The mass fraction of NaCl in the brine solution is 4%; the volume of the brine is 80 mL; and the mass concentration of lime in the brine is 0.04%.

[0049] After obtaining the de-astringent olive leaves, the de-astringent olive leaves are mixed with pure water in a material-liquid mass ratio of 1:8, and then the mixture is squeezed and crushed by a wall breaking machine to obtain olive leaf juice.

[0050] The compound enzyme was added to the olive leaf juice for enzymolysis at a temperature of 60° C. for 2 h. The amount of compound enzyme added was 0.5 g.

[0051] The composition of the compound enzyme is: papain, pectinase, cellulase and neutral protease are mixed in a mass ratio of 1:1:2:1, wherein the enzymatic activity of papain is 2400U / mg; the enzymatic activity of pectinase is 500U / g; the enzymatic activity of cellulase is 10000U / g; and the enzymatic activity of neutral protease is 70U / mg. Papain, pectinase, cellulase and neutral protease are preferably purchased from Nanning Dongheng Huadao Biotechnology Co., Ltd.

[0052] After the enzymatic hydrolysis is completed, the olive leaf enzymatic hydrolysate is placed in a microwave ultrasonic instrument for ultrasonic treatment; the ultrasonic treatment temperature is 50°C, the ultrasonic treatment power is 800W, and the ultrasonic treatment time is 30 minutes. After the ultrasonic treatment is completed, the supernatant is taken to obtain the olive leaf water extract.

[0053] Example 1-2

[0054] A method for preparing an olive leaf water extract comprises the following steps:

[0055] The olive leaves come from olive trees planted in Longnan, Gansu Province. Fresh green leaves picked in March are selected. 20g of fresh green olive leaves are rinsed with clean water several times, and then soaked in pure water with greening agent added for 1 hour to soften the olive leaves.

[0056] The greening agent is greening agent component 2, and the mass ratio of ascorbic acid, citric acid and sodium bicarbonate in the greening agent component 2 is 6:4:1. The preparation method of the greening agent pure water is: add 0.6g ascorbic acid, 0.4g citric acid and 0.1g sodium bicarbonate into 180mL pure water.

[0057] The olive leaves soaked in the above step are immersed in salt water with lime for 3.5 hours to remove bitterness and astringency, and then washed with purified water and dried to obtain de-astringent olive leaves;

[0058] The mass fraction of NaCl in the brine solution is 5%; the volume of the brine is 180 mL; and the mass concentration of lime in the brine is 0.05%.

[0059] After obtaining the de-astringent olive leaves, the de-astringent olive leaves are mixed with pure water in a material-liquid mass ratio of 1:9, and then the mixture is squeezed and crushed by a wall breaking machine to obtain olive leaf juice.

[0060] The compound enzyme was added to the olive leaf juice for enzymolysis at a temperature of 60° C. for 3 h. The amount of compound enzyme added was 1.25 g.

[0061] The composition of the compound enzyme is: papain, pectinase, cellulase and neutral protease are mixed in a mass ratio of 1:1:2:1, wherein the enzymatic activity of papain is 2400U / mg; the enzymatic activity of pectinase is 500U / g; the enzymatic activity of cellulase is 10000U / g; and the enzymatic activity of neutral protease is 70U / mg. Papain, pectinase, cellulase and neutral protease are preferably purchased from Nanning Dongheng Huadao Biotechnology Co., Ltd.

[0062] After the enzymatic hydrolysis is completed, the olive leaf enzymatic hydrolysate is placed in a microwave ultrasonic instrument for ultrasonic treatment; the ultrasonic treatment temperature is 50°C, the ultrasonic treatment power is 800W, and the ultrasonic treatment time is 30min. After the ultrasonic treatment is completed, the supernatant is taken to obtain the olive leaf water extract as shown in FIG. Figure 6 shown.

[0063] Examples 1-3

[0064] A method for preparing an olive leaf water extract comprises the following steps:

[0065] The olive leaves come from olive trees planted in Longnan, Gansu Province. Fresh green leaves picked in March are selected. 30g of fresh green olive leaves are rinsed with clean water several times, and then soaked in pure water with greening agent added for 1 hour to soften the olive leaves.

[0066] The preparation method of the greening agent pure water is as follows: 2.7 g of ascorbic acid, 1.5 g of citric acid and 0.3 g of sodium bicarbonate are added into 300 mL of pure water.

[0067] The olive leaves soaked in the above step are immersed in salt water with lime for 4 hours to remove bitterness and astringency, and then washed with purified water and dried to obtain de-astringent olive leaves;

[0068] The mass fraction of NaCl in the brine solution is 8%; the volume of the brine is 300 mL; and the mass concentration of lime in the brine is 0.05%.

[0069] After obtaining the de-astringent olive leaves, the de-astringent olive leaves are mixed with purified water at a material-liquid mass ratio of 1:10, and then the mixture is squeezed and crushed with a wall breaking machine to obtain olive leaf juice.

[0070] The compound enzyme was added to the olive leaf juice for enzymolysis at a temperature of 60° C. for 4 h. The amount of compound enzyme added was 2 g.

[0071] The composition of the compound enzyme is: papain, pectinase, cellulase and neutral protease are mixed in a mass ratio of 1:1:2:1, wherein the enzymatic activity of papain is 2400U / mg; the enzymatic activity of pectinase is 500U / g; the enzymatic activity of cellulase is 10000U / g; and the enzymatic activity of neutral protease is 70U / mg. Papain, pectinase, cellulase and neutral protease are preferably purchased from Nanning Dongheng Huadao Biotechnology Co., Ltd.

[0072] After the enzymatic hydrolysis is completed, the olive leaf enzymatic hydrolysate is placed in a microwave ultrasonic instrument for ultrasonic treatment; the ultrasonic treatment temperature is 50°C, the ultrasonic treatment power is 800W, and the ultrasonic treatment time is 30 minutes. After the ultrasonic treatment is completed, the supernatant is taken to obtain the olive leaf water extract.

[0073] Comparative Example 1-1

[0074] The preparation method of olive leaf water extract is the same as that of Example 1-3, except that the pectinase in the compound enzyme is replaced with the same mass of saccharifying enzyme, and the enzyme activity of the saccharifying enzyme is 500 U / g.

[0075] Comparative Example 1-2

[0076] The preparation method of olive leaf water extract is the same as that of Example 1-3, except that the papain in the compound enzyme is replaced with alkaline protease of the same mass, and the enzyme activity of the alkaline protease is 2400 U / mg.

[0077] Comparative Examples 1-3

[0078] The preparation method of olive leaf water extract is the same as that of Example 1-3, except that the papain in the composite enzyme is replaced with an equal mass of immobilized glucose isomerase, and the enzyme activity of the immobilized glucose isomerase is 2400 U / mg.

[0079] Comparative Examples 1-4

[0080] The preparation method of olive leaf water extract is the same as that of Example 1-3, except that after obtaining the olive leaf juice, no composite enzyme is added, and the olive leaf juice is directly ultrasonically extracted for 4 hours at 60° C., and the ultrasonic power is 600 W. After the ultrasonic extraction is completed, the ultrasonic extract is filtered and the supernatant is taken to obtain the olive leaf water extract to obtain the conventional olive leaf water extract.

[0081] Comparative Examples 1-5

[0082] The preparation method of olive leaf water extract is the same as that of comparative examples 1-3 of the embodiment, except that the de-astringent olive leaves and ethanol are mixed in a material-liquid mass ratio of 1:10 and then crushed by a wall breaker to obtain olive leaf juice, which is then subjected to a subsequent extraction process.

[0083] Application Example 1

[0084] According to the preparation method of the olive leaf extract of Examples 1-3, olive leaf extracts were prepared respectively. The difference from Examples 1-3 is that the amount of olive leaf is 1g, and the olive leaf extract 1 with 1mg / L of olive leaf extract is prepared; the amount of olive leaf is 2g, and the olive leaf extract 2 with 2mg / L of olive leaf extract is prepared; the amount of olive leaf is 3g, and the olive leaf extract 3 with 3mg / L of olive leaf extract is prepared; the amount of olive leaf is 4g, and the olive leaf extract 4 with 4mg / L of olive leaf extract is prepared; the amount of olive leaf is 5g, and the olive leaf extract 5 with 5mg / L of olive leaf extract is prepared.

[0085] According to the preparation methods of the olive leaf extracts of Comparative Examples 1-4, olive leaf extracts were prepared respectively. The difference from Comparative Examples 1-4 is that the amount of olive leaf is 1 g, and the olive leaf extract 1 with 1 mg / L of olive leaf extract is prepared; the amount of olive leaf is 2 g, and the olive leaf extract 2 with 2 mg / L of olive leaf extract is prepared; the amount of olive leaf is 3 g, and the olive leaf extract 3 with 3 mg / L of olive leaf extract is prepared; the amount of olive leaf is 4 g, and the olive leaf extract 4 with 4 mg / L of olive leaf extract is prepared; the amount of olive leaf is 5 g, and the olive leaf extract 5 with 5 mg / L of olive leaf extract is prepared.

[0086] According to the preparation methods of the olive leaf extracts of Comparative Examples 1-5, olive leaf extracts were prepared respectively. The difference from Comparative Examples 1-5 is that the amount of olive leaf is 1g, and the olive leaf extract 1 with 1mg / L of olive leaf extract is prepared; the amount of olive leaf is 2g, and the olive leaf extract 2 with 2mg / L of olive leaf extract is prepared; the amount of olive leaf is 3g, and the olive leaf extract 3 with 3mg / L of olive leaf extract is prepared; the amount of olive leaf is 4g, and the olive leaf extract 4 with 4mg / L of olive leaf extract is prepared; the amount of olive leaf is 5g, and the olive leaf extract 5 with 5mg / L of olive leaf extract is prepared.

[0087] The free radical scavenging rate of the olive leaf extract prepared above was tested according to the "DPPH and ABTS method for the determination of the antioxidant activity of polypeptides" in GB / T39100-2020. Figure 1 shown.

[0088] Depend on Figure 1 It can be seen that the scavenging rate of DPPH free radicals of olive leaf enzymatic extracts is better than that of the extracts of the other two solvents.

[0089] Comparative Examples 1-6

[0090] The preparation method of the olive leaf water extract is the same as that of Example 1-2, except that the greening agent is different. The preparation method of the greening agent pure water for olive leaf soaking is: add 0.2g of ascorbic acid, 0.6g of citric acid, and 0.1g of sodium bicarbonate to 180mL of pure water. (The mass ratio of greening agent component 3 is ascorbic acid, citric acid and sodium bicarbonate is 2:6:1).

[0091] Comparative Examples 1-7

[0092] The preparation method of olive leaf water extract is the same as that of Example 1-2, except that the greening agent component is greening agent component 4, and the greening agent is only ascorbic acid. The preparation method of greening agent pure water for olive leaf soaking is: add 0.9 g of ascorbic acid to 180 mL of pure water. (Greening agent component 4 is ascorbic acid).

[0093] Comparative Examples 1-8

[0094] The preparation method of olive leaf water extract is the same as that of Example 1-2, except that the greening agent component is greening agent component 5, the mass ratio of ascorbic acid and citric acid of the greening agent component is 3:2, and the preparation method of greening agent pure water for olive leaf soaking is: add 0.6g of ascorbic acid and 0.4g of citric acid to 180mL of pure water. (The mass ratio of ascorbic acid to citric acid is 3:2).

[0095] Comparative Examples 1-9

[0096] The preparation method of the olive leaf water extract is the same as that of Example 1-2, except that in the greening agent soaking step, an equal volume of water is used to replace the greening agent to treat the olive leaves.

[0097] Application Example 2

[0098] During the preparation of the olive leaf water extracts of Examples 1-2 and Comparative Examples 1-9, the olive leaves were photographed before the leaves were mixed with purified water for juicing. The photographic results of Examples 1-2 are as follows: Figure 2 As shown, the photographing results of Comparative Examples 1-9 are as follows Figure 3 shown.

[0099] Depend on Figure 2 and Figure 3 It can be seen that if the greening agent is not added, the chlorophyll is easily accelerated to become pheophytin during the extraction process, so that the color of the olive leaf water extract deteriorates, and changes from green to brown to varying degrees. In the process of preparing the olive leaf water extract, the olive leaf water extract is soaked in a greening agent to keep the leaf color from being significantly deteriorated.

[0100] Application Example 3

[0101] The chlorophyll content in the olive leaf water extract prepared in Example 1-1 and Example 1-2 and Comparative Examples 1-6 to Comparative Examples 1-8 was detected, and the detection method was referred to "NY / T 3082-2017 Spectrophotometric method for determination of chlorophyll content in fruits, vegetables and their products". The chlorophyll content in the olive leaf water extract prepared in Example 1-1 and Example 1-2 and Comparative Examples 6 to Comparative Examples 8 is as follows: Figure 4 As shown. Among them, the detection results of chlorophyll in the olive leaf water extract prepared in Example 1-1 corresponding to the green protectant component 1; the detection results of chlorophyll in the olive leaf water extract prepared in Example 1-2 corresponding to the green protectant component 2; the detection results of chlorophyll in the olive leaf water extract prepared in Comparative Examples 1-6 corresponding to the green protectant component 3; the detection results of chlorophyll in the olive leaf water extract prepared in Comparative Examples 1-7 corresponding to the green protectant component 4; the detection results of chlorophyll in the olive leaf water extract prepared in Comparative Examples 1-8 corresponding to the green protectant component 5.

[0102] Depend on Figure 4 It can be seen that the chlorophyll content in the olive leaf water extract varies greatly when different components of green protectants are used. Compared with green protectant component 1 and green protectant component 2, the chlorophyll content in the olive leaf extract of green protectant component 3, green protectant component 4 and green protectant component 5 is significantly lower, and the optimal green protectant component ratio is the mass ratio of ascorbic acid, citric acid and sodium bicarbonate is 6:4:1.

[0103] Application Example 4

[0104] The active ingredients of the olive leaf water extracts prepared in Examples 1-2 to 1-3 and Comparative Examples 1-4 and 1-5 were detected by ultra-high performance liquid chromatography tandem mass spectrometry. The test results are shown in Figure 5 Among them, 1 is oleuropein, 2 is hydroxytyrosol, 3 is verbascoside, 4 is olibanum glucoside, and 5 is gallic acid.

[0105] Depend on Figure 5 It can be seen that the main active ingredients in the olive leaf water extract 5 prepared in Examples 1-2 to 1-3 are significantly higher than those in the comparative example, especially in Example 1-3, the content of active ingredients is the highest.

[0106] In summary, the preparation method of the olive leaf water extract provided by the present invention can fully extract the active ingredients in the olive leaves. The present invention extracts the active ingredients in the olive leaves so that the active ingredients in the olive leaves are easily absorbed by the body.

[0107] Although the above embodiment describes the present invention in detail, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A method for preparing an olive leaf aqueous extract, characterized in that: The following steps are involved: The olive leaves are softened and then debittered to obtain de-astringent olive leaves; The de-astringent olive leaves are crushed and then enzymolyzed with a composite enzyme to obtain an olive leaf hydrolyzate; the composite enzyme comprises papain, pectinase, cellulase and neutral protease; the mass ratio of papain, pectinase, cellulase and neutral protease in the composite enzyme is 1:1:2:1; The enzymatic activity of the papain is 2400U / mg; the enzymatic activity of the pectinase is 500U / g; the enzymatic activity of the cellulase is 10000U / g; the enzymatic activity of the neutral protease is 70U / mg; The mass ratio of the olive leaf to the complex enzyme is (10-50): (0.5-1.5); The enzymatic hydrolysis temperature is 40°C to 70°C; the enzymatic hydrolysis time is 2 to 4 hours; subjecting the olive leaf enzymatic hydrolysate to ultrasonic treatment to obtain an olive leaf water extract; The softening process includes soaking the olive leaves in an aqueous solution of a green preserving agent; the soaking time is 0.5 to 2 hours; The greening agent comprises ascorbic acid, citric acid and sodium bicarbonate; the mass ratio of ascorbic acid, citric acid and sodium bicarbonate in the greening agent is (3-9):(3-5):1; In the greening agent aqueous solution, the mass ratio of the greening agent to water is (5-50):(500-4000); The debittering agent includes a brine solution containing lime; the mass concentration of NaCl in the brine solution containing lime is 4% to 8%; the mass concentration of lime in the brine solution containing lime is 0.04% to 0.06%; The debittering time is 3 to 4 hours.

2. The preparation method according to claim 1, characterized in that: The mass ratio of the olive leaf to the complex enzyme is (10-30): (0.5-1.5).

3. The preparation method according to claim 1, characterized in that: The mass ratio of the olive leaves to the complex enzyme is 20:

1.

4. The preparation method according to claim 1, characterized in that: The temperature of the enzymolysis is 50°C to 70°C; the time of the enzymolysis is 3 hours.

5. The preparation method according to claim 1, characterized in that: The temperature of the enzymolysis is 60° C.; the time of the enzymolysis is 3 h.

6. The preparation method according to claim 1, characterized in that: The power of the ultrasound is 600-1000W, the time of the ultrasound is 15-45min, and the temperature of the ultrasound is 50°C-70°C.

7. The preparation method according to claim 1, characterized in that: The mass ratio of ascorbic acid, citric acid and sodium bicarbonate in the greening agent is (4-8):(3-5):

1.

8. The preparation method according to claim 1, characterized in that: The mass ratio of ascorbic acid, citric acid and sodium bicarbonate in the greening agent is 6:4:

1.

9. The preparation method according to claim 1, characterized in that: In the greening agent aqueous solution, the mass ratio of the greening agent to water is (7-45):(800-3000).

10. The preparation method according to claim 1, characterized in that: In the greening agent aqueous solution, the mass ratio of the greening agent to water is 11:1800.

11. The preparation method according to claim 1, characterized in that: The mass fraction of NaCl in the lime-containing brine solution is 6%; the mass concentration of lime in the lime-containing brine solution is 0.05%.

12. An olive leaf water extract prepared by the preparation method according to any one of claims 1 to 11.

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