Method for extracting leaves of psidium plants, leaf extract of psidium plants obtained by the same, and physiologically active composition containing leaf extract of psidium plants

Abstract

To provide a method for extracting active ingredients from the leaves of Psidium plants.SOLUTION: A method for extracting leaves of Psidium plants includes: (a) preparing crushed or powdered leaves of Psidium plants; (b) mixing crushed or powdered leaves of Psidium plants with water to prepare a crushed leaf mixture or a leaf powder suspension; (c) adding an enzyme preparation to the crushed leaf mixture or leaf powder suspension to prepare a first mixture, and performing an enzyme treatment process to obtain an enzyme-treated product, the weight ratio of the powdered leaves of Psidium plants to the enzyme preparation being 1:0.01 to 1; and (d) mixing the enzyme-treated product with an alcohol solvent to prepare a second mixture, performing an extraction process, and obtaining an extraction liquid, the extraction liquid containing at least one active ingredient.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The present invention relates to a method for extracting plant materials, and particularly to a method for extracting leaves of Psidium guajava plants, a leaf extract of Psidium plants obtained thereby, and a bioactive composition containing the leaf extract of Psidium plants.

Background Art

[0002] A large amount of cellulose, hemicellulose, protein, etc. are present in plant tissues of agricultural by-products. However, due to these components, the efficiency of extracting active substances from agricultural by-products in the pretreatment and purification processes for reuse is greatly limited, and the concentration of the remaining active substances after purification is not high. Furthermore, the active substance-containing solution obtained as described above tends to form insoluble complexes due to the bonds formed mainly by tannin catechins or gallic acid contained therein during storage, resulting in precipitation and turbidity, thereby affecting the stability and activity of the active substances.

[0003] The cultivation of Psidium guajava (guava) promotes growth and fruiting when pruning of branches and leaves is carried out in a timely manner. A considerable amount of branch and leaf waste is generated during the pruning process, and these discarded Psidium guajava leaves are often subjected to landfill and incineration treatments, potentially leading to CO2 emissions. Currently, since Psidium guajava leaf extract is known to have a high concentration of tannin components, it can be a target for effective recycling and reuse of agricultural by-products and / or agricultural waste.

[0004] Therefore, there is an extremely urgent need for a method that can more effectively reuse or extract agricultural by-products and / or agricultural waste.

Summary of the Invention

Problems to be Solved by the Invention

[0005] Timely pruning of the branches and leaves required for cultivating plants of the genus Psidium produces waste such as branches and leaves, and also results in a large amount of agricultural by-products in terms of agricultural cultivation. The main problem of the present disclosure is to provide a method for effectively recycling or extracting these agricultural by-products and / or agricultural wastes. In addition, the present disclosure also provides a solution to the precipitation phenomenon that is likely to occur during the process of plant extraction.

Means for Solving the Problems

[0006] The present disclosure provides a method for effectively recycling the leaves of plants of the genus Psidium and a product obtained by this method.

[0007] That is, the present disclosure may include, but is not limited to, the following contents.

[0008] [1] A method for extracting the leaves of plants of the genus Psidium (Psidium), comprising: (a) providing crushed pieces or powder of the leaves of plants of the genus Psidium; (b) mixing the crushed pieces or powder of the leaves of the plants of the genus Psidium with water at a weight ratio of 1:7 to 21 to prepare a leaf crushed piece mixture or a leaf powder suspension; (c) adding an enzyme preparation to the leaf crushed piece mixture or the leaf powder suspension to prepare a first mixture, and performing an enzyme treatment step for 0.5 to 24 hours to obtain an enzyme-treated product, wherein the weight ratio of the crushed pieces or powder of the leaves of the plants of the genus Psidium to the enzyme preparation is 1:0.01 to 1, and the formulation of the enzyme preparation includes one of the formulations shown in the following (i) to (iv): (i) containing an enzyme mixture, the enzyme mixture including arabinase, cellulase, β-glucanase, hemicellulase and xylanase, formulation (I) (ii) formulation (II) containing the enzyme mixture and pectinase (iii) Formulation (III) containing tannase (iv) Formulation (IV) containing the enzyme mixture, the pectinase and the tannase (d) A step of mixing the enzyme-treated product and an alcohol solvent at a weight ratio of 1:0.1 to 2.5 to form a second mixture, and performing an extraction step to obtain an extract as an extract of the leaves of the Psidium plant, wherein the extract contains at least one active ingredient An extraction method for the leaves of a Psidium plant containing the above

[0009] [2] The extraction method for the leaves of the Psidium plant according to [1], wherein the Psidium plant includes Psidium guajava, Psidium littorale or Psidium acutangulum

[0010] [3] The extraction method for the leaves of the Psidium plant according to [1], wherein the Psidium plant is Psidium guajava

[0011] [4] The at least one active ingredient includes the following components The extraction method for the leaves of the Psidium plant according to [1], wherein at least one of polysaccharides and polyphenols is included

[0012] [5] The polysaccharides include the following polysaccharides At least one of glucan, fructan, xylan and glucomannan The extraction method for the leaves of the Psidium plant according to [4], wherein at least one of them is included

[0013] [6] The polyphenols include the following polyphenols The extraction method for the leaves of the Psidium plant according to [4], wherein at least one of phenolic acids, flavonoids, stilbenoids and lignans is included

[0014] [7] The phenolic acids include the following phenols: at least one of ellagic acid, gallic acid, and quinic acid is included, and [8] The flavonoids include the following flavonoids: at least one of quercetin and kaempferol is included, the method for extracting leaves of the genus Banjirow described in [6].

[0015] [8] The size of the crushed pieces or powder of the leaves of the genus Banjirow is 0.05 - 0.5 mm, the method for extracting leaves of the genus Banjirow described in [1].

[0016] [9] In the step (b), the pH value of the leaf crushed piece mixture or leaf powder suspension is adjusted to be 4.5 - 5.5, the method for extracting leaves of the genus Banjirow described in [1].

[0017]

[10] In the step (c), the enzyme treatment step is carried out at 35 - 55 °C, the method for extracting leaves of the genus Banjirow described in [1].

[0018]

[11] In the step (c), the weight ratio of the crushed pieces or powder of the leaves of the genus Banjirow to the enzyme preparation is 1:0.1 - 0.3, the method for extracting leaves of the genus Banjirow described in [1].

[0019]

[12] In the compounding formula (II), the weight ratio of the enzyme mixture to the pectinase is 1:0.5 - 2, the method for extracting leaves of the genus Banjirow described in [1].

[0020]

[13] In the compounding formula (IV), the weight ratio of the enzyme mixture, the pectinase, and the tannase is 1:0.5 - 2:0.5 - 2, the method for extracting leaves of the genus Banjirow described in [1].

[0021]

[14] In the step (c), the formulation of the enzyme preparation is the formulation (I), and the weight ratio of the crushed pieces or powder of the leaves of the plant of the genus Banziro to the enzyme preparation is 1:0.1. The method for extracting the leaves of the plant of the genus Banziro according to [1].

[0022]

[15] In the step (c), the formulation of the enzyme preparation is the formulation (II), the weight ratio of the crushed pieces or powder of the leaves of the plant of the genus Banziro to the enzyme preparation is 1:0.2, and in the formulation (II), the weight ratio of the enzyme mixture to the pectinase is 1:1. The method for extracting the leaves of the plant of the genus Banziro according to [1].

[0023]

[16] In the step (c), the formulation of the enzyme preparation is the formulation (III), and the weight ratio of the crushed pieces or powder of the leaves of the plant of the genus Banziro to the enzyme preparation is 1:0.1. The method for extracting the leaves of the plant of the genus Banziro according to [1].

[0024]

[17] During the step (c), the formulation of the enzyme preparation is the formulation (IV), the weight ratio of the crushed pieces or powder of the leaves of the plant of the genus Banziro to the enzyme preparation is 1:0.3, and in the formulation (IV), the weight ratio of the enzyme mixture, the pectinase and the tannase is 1:1:1. The method for extracting the leaves of the plant of the genus Banziro according to [1].

[0025]

[18] In the step (d), in the second mixture, the final concentration of the alcohol solvent is 20 - 50%. The method for extracting the leaves of the plant of the genus Banziro according to [1].

[0026]

[19] In the step (d), the alcohol solvent includes methanol, ethanol, ethylene glycol, propanol or isopropanol. The method for extracting the leaves of the plant of the genus Banziro according to [1].

[0027]

[20] In the step (d), the extraction process is carried out at 50 to 70 °C for 0.5 to 24 hours, the extraction method of the leaves of the genus Bangirola described in [1].

[0028]

[21] After the step (d), (e) A step of performing a filtration process on the extract to obtain a filtrate, the filtrate containing the at least one active ingredient, the extraction method of the leaves of the genus Bangirola described in [1] further including this step.

[0029]

[22] After the step (d), (f) A step of performing a centrifugation process on the filtrate to obtain a supernatant, the supernatant containing the at least one active ingredient, the extraction method of the leaves of the genus Bangirola described in

[21] further including this step.

[0030]

[23] After the step (f), (g) A step of performing a drying process on the supernatant to form an extract powder, the extract powder containing the at least one active ingredient, the extraction method of the leaves of the genus Bangirola described in

[22] further including this step.

[0031]

[24] A leaf extract of the genus Bangirola containing at least one physiologically active ingredient and obtained by the extraction method of the leaves of the genus Bangirola described in any one of [1] to

[23] .

[0032]

[25] A physiologically active composition containing the leaf extract of the genus Bangirola described in

[24] .

[0033]

[26] A pharmaceutical composition, and further containing a pharmaceutically acceptable carrier or salt, the physiologically active composition described in

[25] .

[0034]

[27] A health supplement food composition, and further containing a food-acceptable additive, the physiologically active composition described in

[25] .

[0035] The bioactive composition according to

[25] , which is a skin care composition and further comprises a pharmaceutically acceptable carrier.

[0036] In order to make the above and other objects, features, and advantages of the present disclosure clearer and easier to understand, preferred embodiments will be given below and described in detail while corresponding to the drawings.

Brief Description of the Drawings

[0037]

Figure 1

Figure 2

Figure 3

Figure 4

Modes for Carrying Out the Invention

[0038] The present disclosure can provide a method for extracting leaves of plants of the genus Psidium.

[0039] According to the method for extracting leaves of plants of the genus Psidium of the present disclosure, agricultural by-products and / or agricultural wastes obtained after harvesting the economic parts of plants of the genus Psidium can be effectively recycled and utilized, and thereby components with economic value, particularly components with biological activity, can be obtained, enhancing the overall economic value of agricultural crops and further promoting recycling use.

[0040] The extraction method of the leaves of the genus Pandanus in the present disclosure can effectively increase the extraction amount of the active ingredients in the leaves of the genus Pandanus by combining specific enzymatic treatment and solvent extraction, and / or can reduce the ester bonds and depside bonds formed by high-concentration tannins in the extract, thereby improving the extraction efficiency, reducing the occurrence of precipitation, and enhancing the stability.

[0041] Specifically, by using the extraction method of the leaves of the genus Pandanus in the present disclosure, the extraction efficiency of the active ingredients extracted from the leaves of the genus Pandanus can be increased, and agricultural by-products and / or agricultural waste can be effectively utilized.

[0042] The above active ingredients may include, but are not limited to, polysaccharides, polyphenols, etc., or any combination thereof.

[0043] The above polysaccharides may include, but are not limited to, glucan, fructan, xylan, glucomannan, etc., or any combination thereof.

[0044] The above polyphenols may include, but are not limited to, phenolic acids, flavonoids, stilbenoids, lignans, etc., or any combination thereof. Examples of the above phenolic acids may include, but are not limited to, ellagic acid, gallic acid, quinic acid, etc., or any combination thereof. Also, examples of the above flavonoids may include, but are not limited to, quercetin, kaempferol, etc., or any combination thereof.

[0045] In the method for extracting leaves of plants of the genus Psidium according to the present disclosure, there is no particular limitation on the plants of the genus Psidium described above, and any plant of the genus Psidium may be used. Examples of plants of the genus Psidium include, but are not limited to, Psidium guajava, Psidium littorale, Psidium acutangulum, etc. In one embodiment, the plant of the genus Psidium in the method for extracting leaves of plants of the genus Psidium according to the present disclosure is Psidium guajava.

[0046] The method for extracting leaves of plants of the genus Psidium according to the present disclosure described above may include, but is not limited to, the following steps.

[0047] First, prepare crushed pieces or powder of the leaves of plants of the genus Psidium. There is no particular limitation on the method for obtaining the crushed pieces or powder of the leaves of plants of the genus Psidium, and any method that can obtain the crushed pieces or powder of the leaves of plants of the genus Psidium may be used. In one embodiment, the leaves or dried leaves can be crushed or pulverized into crushed pieces or powder, but it is not limited thereto. Also, there is no particular limitation on the size of the crushed pieces or powder of the leaves of plants of the genus Psidium, and any size that facilitates the extraction of active ingredients in the subsequent leaves may be used. In one embodiment, the size of the crushed pieces or powder of the leaves of plants of the genus Psidium is about 0.05 to 0.5 mm, for example, about 0.1 to 0.5 mm, about 0.2 to 0.4 mm, about 0.1 mm, about 0.2 mm, about 0.25 mm, about 0.3 mm, about 0.4 mm, about 0.5 mm, etc., but it is not limited thereto. In a specific embodiment, the size of the crushed pieces or powder of the leaves of plants of the genus Psidium may be 0.25 mm.

[0048] In the present disclosure, "about" represents a range of ±10% to ±1% of the stated amount, for example, a range of ±10%, a range of ±5%, a range of ±1%, etc., but it is not limited thereto.

[0049] Next, the crushed pieces or powder of the leaves of the above-mentioned *Banjilou* plants are mixed with water to prepare a leaf crushed piece mixed solution or a leaf powder suspension. The weight ratio of the crushed pieces or powder of the leaves of the above-mentioned *Banjilou* plants to water may be about 1:7 to 21, for example, about 1:8 to 20, about 1:10 to 18, about 1:11 to 15, about 1:7 to 14, about 1:7, about 1:8, about 1:9, about 1:10, about 1:11, about 1:12, about 1:13, about 1:14, about 1:15, about 1:16, about 1:17, about 1:18, about 1:19, about 1:20, about 1:21, but is not limited thereto. In one embodiment, the weight ratio of the crushed pieces or powder of the leaves of the above-mentioned *Banjilou* plants to water may be about 1:14.

[0050] Optionally, the pH value of the above-mentioned leaf crushed piece mixed solution or leaf powder suspension can be adjusted. For example, the pH value of the above-mentioned leaf crushed piece mixed solution or leaf powder suspension can be adjusted to about 4.5 to 5.5, for example, about 4.5, about 4.6, about 4.7, about 4.8, about 4.9, about 5.0, about 5.1, about 5.2, about 5.3, about 5.4, about 5.5, etc., but is not limited thereto. In one embodiment, the pH value of the above-mentioned leaf crushed piece mixed solution or leaf powder suspension can be adjusted to about 5.0.

[0051] In the method for extracting the leaves of the *Banjilou* plants of the present disclosure, after the crushed pieces or powder of the leaves of the above-mentioned *Banjilou* plants are mixed with water to prepare a leaf crushed piece mixed solution or a leaf powder suspension, an enzyme preparation can be added to the above-mentioned leaf crushed piece mixed solution or leaf powder suspension to prepare a first mixture, and an enzyme treatment step can be performed to obtain an enzyme-treated product.

[0052] The weight ratio of the crushed pieces or powder of the leaves of the above-mentioned *Banilla* plant to the above-mentioned enzyme preparation may be about 1:0.01 to 1, for example, about 1:0.02 to 0.9, about 1:0.05 to 0.8, about 1:0.1 to 0.7, about 1:0.1 to 0.3, about 1:0.2 to 0.6, about 1:0.3 to 0.5, about 1:0.01, about 1:0.05, about 1:0.1, about 1:0.2, about 1:0.3, about 1:0.4, about 1:0.5, about 1:0.6, about 1:0.7, about 1:0.8, about 1:0.9, about 1:1, etc., but is not limited thereto. In one embodiment, the weight ratio of the crushed pieces or powder of the leaves of the above-mentioned *Banilla* plant to the above-mentioned enzyme preparation may be about 1:0.1. In another embodiment, the weight ratio of the crushed pieces or powder of the leaves of the above-mentioned *Banilla* plant to the above-mentioned enzyme preparation may be about 1:0.2. In yet another embodiment, the weight ratio of the crushed pieces or powder of the leaves of the above-mentioned *Banilla* plant to the above-mentioned enzyme preparation may be about 1:0.3.

[0053] The formulation of the above-mentioned enzyme preparation may include, but is not limited to, one of the following formulations (I) to (IV).

[0054] Formulation (I) may contain an enzyme mixture. The above-mentioned enzyme mixture may contain arabinase, cellulase, β-glucanase, hemicellulase, and xylanase. Examples of the above-mentioned enzyme mixture may include, but are not limited to, commercially available Viscozyme® L.

[0055] The composition formulation (II) may contain the enzyme mixture and pectinase of the above composition formulation (I). In the above composition formulation (II), the weight ratio of the enzyme mixture to the pectinase may be about 1:0.5 to 2, such as about 1:0.5 to 1.8, about 1:0.6 to 1.7, about 1:0.5, about 1:0.6, about 1:0.7, about 1:0.8, about 1:0.9, about 1:1, about 1:1.1, about 1:1.2, about 1:1.3, about 1:1.4, about 1:1.5, about 1:1.6, about 1:1.7, about 1:1.8, about 1:1.9, about 1:2, etc., but is not limited thereto. In one embodiment, in the above composition formulation (II), the weight ratio of the enzyme mixture to the pectinase may be about 1:1.

[0056] The composition formulation (III) may contain tannase.

[0057] The composition formulation (IV) may contain the enzyme mixture, pectinase and tannase of the above composition formulation (I). In the above composition formulation (IV), the weight ratio of the enzyme mixture, the pectinase and the tannase may be about 1:0.5 to 2:0.5 to 2, such as about 1:0.5 to 1.8:0.5 to 1.8, about 1:0.6 to 1.7:0.6 to 1.7, about 1:0.5:0.5, about 1:0.5:1, about 1:1:0.5, about 1:0.6:0.6, about 1:0.7:0.7, about 1:0.8:0.8, about 1:0.9:0.9, about 1:1:1, about 1:1.1:1.1, about 1:1.2:1.2, about 1:1.3:1.3, about 1:1.4:1.4, about 1:1.5:1.5, about 1:1:1.5, about 1:1.5:1, about 1:1.6:1.6, about 1:1.7:1.7, about 1:1.8:1.8, about 1:1.9:1.9, about 1:1:2, about 1:2:2, about 1:2:1, etc., but is not limited thereto. In one embodiment, in the above composition formulation (IV), the weight ratio of the enzyme mixture, the pectinase and the tannase may be about 1:1:1.

[0058] In one embodiment, in the step of adding an enzyme preparation to the above leaf crushed fragment mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, the formulation of the enzyme preparation may be the above formulation (I), and the weight ratio of the crushed fragments or powder of the leaves of the plant of the genus Pandanus to the enzyme preparation may be about 1:0.1.

[0059] In another embodiment, in the step of adding an enzyme preparation to the above leaf crushed fragment mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, the formulation of the enzyme preparation may be the above formulation (II), the weight ratio of the crushed fragments or powder of the leaves of the plant of the genus Pandanus to the enzyme preparation may be about 1:0.2, and in the above formulation (II), the weight ratio of the enzyme mixture to the pectinase may be about 1:1.

[0060] In yet another embodiment, in the step of adding an enzyme preparation to the above leaf crushed fragment mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, the formulation of the enzyme preparation may be the above formulation (III), and the weight ratio of the crushed fragments or powder of the leaves of the plant of the genus Pandanus to the enzyme preparation may be about 1:0.1.

[0061] In still another embodiment, in the step of adding an enzyme preparation to the above leaf crushed fragment mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, the formulation of the enzyme preparation may be the above formulation (IV), the weight ratio of the crushed fragments or powder of the leaves of the plant of the genus Pandanus to the enzyme preparation may be about 1:0.3, and in the above formulation (IV), the weight ratio of the enzyme mixture, the pectinase, and the tannase may be about 1:1:1.

[0062] In the step of adding an enzyme preparation to the above-mentioned leaf crushed piece mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, there is no particular limitation on the temperature at which the enzyme treatment step is performed, as long as the enzyme preparation can have the required enzyme activity at the adopted temperature. In one embodiment, the enzyme treatment step can be performed at about 35 to 55 °C, for example, about 36 to 54 °C, about 37 to 53 °C, about 38 to 50 °C, about 39 to 51 °C, about 40 to 50 °C, about 35 °C, about 36 °C, about 37 °C, about 37.5 °C, about 38 °C, about 39 °C, about 40 °C, about 41 °C, about 42 °C, about 43 °C, about 44 °C, about 45 °C, about 46 °C, about 47 °C, about 48 °C, about 49 °C, about 50 °C, about 51 °C, about 52 °C, about 53 °C, about 54 °C, about 55 °C, etc., but is not limited thereto. In a specific embodiment, in the step of adding an enzyme preparation to the above-mentioned leaf crushed piece mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, the enzyme treatment step can be performed at about 45 °C or lower.

[0063] In the step of adding an enzyme preparation to the above-mentioned leaf crushed piece mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, there is no particular limitation on the time for which the enzyme treatment step proceeds, as long as it is a time for which the enzyme treatment step can proceed sufficiently. In one embodiment, the enzyme treatment step can be carried out for about 0.5 to 24 hours, for example, about 0.5 to 21 hours, about 1 to 18 hours, about 2 to 18 hours, about 2.5 to 15 hours, about 3 to 15 hours, about 4 to 12 hours, about 5 to 10 hours, about 0.5 hour, about 1 hour, about 1.5 hours, about 2 hours, about 2.5 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 8 hours, about 9 hours, about 10 hours, about 12 hours, about 14 hours, about 15 hours, about 16 hours, about 18 hours, about 20 hours, about 21 hours, about 22 hours, about 24 hours, but is not limited thereto. In a specific embodiment, in the step of adding an enzyme preparation to the above-mentioned leaf crushed piece mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, the enzyme treatment step can be carried out for about 1 hour.

[0064] Finally, after the step of adding the enzyme preparation to the above leaf crushed fragment mixture or leaf powder suspension to form a first mixture and performing an enzyme treatment step to obtain an enzyme-treated product, the enzyme-treated product and an alcohol solvent are mixed to form a second mixture, and an extraction step is performed to obtain an extract as an extract of the leaves of the genus Banjiro. The above extract may contain at least one active ingredient.

[0065] In the above step of mixing the enzyme-treated product and an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of the genus Banjiro, the weight ratio of the enzyme-treated product to the alcohol solvent is about 1:0.1 to 2.5, for example, about 1:0.15 to 2.2, about 1:0.2 to 2.1, about 1:0.3 to 2, about 1:0.4 to 1.8, about 1:0.45 to 1.5, about 1:0.5 to 1, about 1:0.6 to 0.8, about 1:0.1, about 1:0.2, about 1:0.3, about 1:0.4, about 1:0.41, about 1:0.42, about 1:0.43, about 1:0.44, about 1:0.45, about 1:0.5, about 1:0.6, about 1:0.7, about 1:0.8, about 1:0.9, about 1:1, about 1:1.5, about 1:2, about 1:2.5, etc., but is not limited thereto. In one embodiment, the weight ratio of the enzyme-treated product to the alcohol solvent may be about 1:0.43. In another embodiment, the weight ratio of the enzyme-treated product to the alcohol solvent may be about 1:0.42.

[0066] Also, in the above step of mixing the enzyme-treated product and an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of the genus Banjiro, in the above second mixture, the final concentration of the alcohol solvent is about 20 to 50%, for example, about 20 to 48%, about 22 to 47%, about 25 to 46%, about 30 to 45%, about 35 to 40%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, etc., but is not limited thereto. In one embodiment, in the above step of mixing the enzyme-treated product and an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of the genus Banjiro, in the above second mixture, the final concentration of the alcohol solvent may be about 30%.

[0067] Furthermore, in the step of mixing the enzyme-treated product with an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of a Banjiro plant, examples of the alcohol solvent include, but are not limited to, methanol, ethanol, ethylene glycol, propanol, isopropanol, etc., or any combination thereof. In one embodiment, in the step of mixing the enzyme-treated product with an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of a Banjiro plant, the alcohol solvent may be ethanol.

[0068] In the step of mixing the enzyme-treated product with an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of a Banjiro plant, there is no particular limitation on the temperature at which the extraction step is performed, as long as it can extract the active ingredient. In one embodiment, the extraction step can be performed at about 50 to 70 °C, for example, about 52 to 68 °C, about 53 to 67 °C, about 54 to 66 °C, about 55 to 65 °C, about 56 to 64 °C, about 57 to 63 °C, about 50 °C, about 51 °C, about 52 °C, about 53 °C, about 54 °C, about 55 °C, about 56 °C, about 57 °C, about 58 °C, about 59 °C, about 60 °C, about 61 °C, about 62 °C, about 63 °C, about 64 °C, about 65 °C, about 66 °C, about 67 °C, about 68 °C, about 69 °C, about 70 °C, etc., but is not limited thereto. In a specific embodiment, in the step of mixing the enzyme-treated product with an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of a Banjiro plant, the extraction step can be performed at about 60 °C.

[0069] In the step of mixing the enzyme-treated product with an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of a Banjiro plant, the time for performing the extraction step is not particularly limited, and any time that allows the extraction step to proceed sufficiently may be used. In one embodiment, the extraction step may be carried out for about 0.5 to 24 hours, for example, about 0.5 to 21 hours, about 1 to 18 hours, about 2 to 18 hours, about 2.5 to 15 hours, about 3 to 15 hours, about 4 to 12 hours, about 5 to 10 hours, about 0.5 hour, about 1 hour, about 1.5 hours, about 2 hours, about 2.5 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 8 hours, about 9 hours, about 10 hours, about 12 hours, about 14 hours, about 15 hours, about 16 hours, about 18 hours, about 20 hours, about 21 hours, about 22 hours, about 24 hours, but is not limited thereto. In a specific embodiment, in the step of mixing the enzyme-treated product with an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of a Banjiro plant, the extraction step can be carried out for about 1 hour.

[0070] In the step of mixing the enzyme-treated product with an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of a Banjiro plant, the at least one active ingredient contained in the obtained extract may include, but is not limited to, polysaccharides, polyphenols, etc., or any combination thereof.

[0071] The polysaccharides may include, but are not limited to, glucan, fructan, xylan, glucomannan, etc., or any combination thereof.

[0072] The above-mentioned polyphenols may include, but are not limited to, phenolic acids, flavonoids, stilbenoids, lignans, etc., or any combination thereof. Examples of the above-mentioned phenolic acids may include, but are not limited to, gallic acid, ellagic acid, caffeic acid, etc., or any combination thereof. Examples of the above-mentioned flavonoids may include, but are not limited to, quercetin, kaempferol, etc., or any combination thereof.

[0073] In one embodiment, in the step of mixing the enzyme-treated product with an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as an extract of the leaves of the genus Pandanus, at least one active ingredient contained in the obtained extract may include polysaccharides, gallic acid, ellagic acid, and quercetin.

[0074] The method for extracting the leaves of the genus Pandanus of the present disclosure may include, but is not limited to, the following steps.

[0075] Also, in one embodiment, the method for extracting the leaves of the genus Pandanus of the present disclosure may further include, in addition to the above steps, a leaf treatment step before the step of preparing the crushed pieces or powder of the leaves of the genus Pandanus to obtain the crushed pieces or powder of the leaves of the genus Pandanus, but is not limited thereto.

[0076] The above leaf treatment step may include, but is not limited to, the following steps.

[0077] First, a drying step is performed on the leaves of the genus Pandanus to obtain dried leaves.

[0078] There is no particular limitation on the method for performing the above drying step, and it can be adjusted according to the requirements and / or the environmental conditions at that time, as long as the leaves can be dried and the active ingredients in the leaves are not adversely affected. In one embodiment, the above drying step can be performed by baking.

[0079] There is no particular limitation on the temperature for performing the above drying process, and it can be adjusted according to the requirements and / or the environmental conditions at that time, as long as it can dry the leaves and does not adversely affect the active ingredients in the leaves. In one embodiment, the above drying process is carried out by baking at about 40 to 60 °C, for example, about 42 to 58 °C, about 43 to 57 °C, about 44 to 56 °C, about 45 to 55 °C, about 46 to 54 °C, about 47 to 53 °C, about 40 °C, about 41 °C, about 42 °C, about 43 °C, about 44 °C, about 45 °C, about 46 °C, about 47 °C, about 48 °C, about 49 °C, about 50 °C, about 51 °C, about 52 °C, about 53 °C, about 54 °C, about 55 °C, about 56 °C, about 57 °C, about 58 °C, about 59 °C, about 60 °C, etc., but it is not limited thereto. In a specific embodiment, the above drying process can be carried out at about 50 °C.

[0080] There is no particular limitation on the time for performing the above drying process, and it can be adjusted according to the requirements and / or the environmental conditions at that time, as long as it can dry the leaves and does not adversely affect the active ingredients in the leaves.

[0081] In one embodiment, the above drying process is carried out on the leaves of the plant of the genus Banjiro until the leaves are dried at 50 °C.

[0082] Also, in the above leaf treatment process, after the step of obtaining dried leaves by performing a drying process on the leaves of the plant of the genus Banjiro, a crushing or pulverizing process is performed on the dried leaves to obtain crushed pieces or powder of the leaves of the plant of the genus Banjiro.

[0083] There is no particular limitation on the method for performing the above crushing or pulverizing process, and it can be adjusted according to the requirements and / or the environmental conditions at that time, as long as the dried leaves can be made into crushed pieces or powder.

[0084] Furthermore, in one embodiment, the above leaf treatment process may further include a pretreatment process before the step of obtaining dried leaves by performing a drying process on the leaves of the plant of the genus Banjiro.

[0085] The above pretreatment process is not limited, but may include the following steps.

[0086] First, select leaves with good condition and no signs of withering or rotting from the branches and leaves of the mango, then separate them from the branches, remove the petioles to obtain the selected leaves. Then, wash the selected leaves thoroughly to obtain the washed leaves. The washed leaves can be used as the leaves of the mango plant in the above leaf treatment step.

[0087] In another embodiment, in addition to the above steps, the method for extracting the leaves of the mango plant of the present disclosure further includes, after the step of mixing the enzyme-treated product and an alcohol solvent to form a second mixture and performing an extraction step to obtain an extract as the leaf extract of the mango plant, performing at least one filtration step on the extract to obtain a filtrate. The filtrate may contain at least one active ingredient, but is not limited thereto. There is no particular limitation on the method adopted in the at least one filtration step, as long as it can remove the solids in the extract. There is also no particular limitation on the number of times of performing the filtration step, which can be determined according to the necessity and / or the environmental conditions at that time. When the number of filtration steps is more than one, the methods adopted in each filtration step may be the same or different. For example, filter papers with different pore sizes can be used to perform the at least one filtration step on the extract.

[0088] In this embodiment, after the step of performing at least one filtration step on the extract to obtain a filtrate, the method may further include performing a centrifugation step on the filtrate to obtain a supernatant. The supernatant may contain at least one active ingredient.

[0089] Furthermore, in this embodiment, after the step of centrifuging the filtrate to obtain a supernatant, a step of drying the supernatant to produce an extract powder may be further included, and the extract powder may contain the at least one active ingredient. There is no particular limitation on the method of performing the drying step, and it can be adjusted according to requirements and / or the environmental conditions at that time, as long as the supernatant can be dried to form an extract powder without adversely affecting the active ingredients therein. In one embodiment, the drying step can be performed by freeze-drying.

[0090] In addition, the present disclosure can also provide a leaf extract of a Saururus plant. The leaf extract of the Saururus plant of the present disclosure may contain at least one physiologically active ingredient, and it may be obtained from any of the above-described extraction methods of the leaves of the Saururus plant of the present disclosure.

[0091] Furthermore, the present disclosure can also provide a physiologically active composition. The physiologically active composition of the present disclosure may include, but is not limited to, any of the above-described leaf extracts of the Saururus plant of the present disclosure.

[0092] In one embodiment, the physiologically active composition of the present disclosure may be a pharmaceutical composition, and may include, but is not limited to, a pharmaceutically acceptable carrier or salt. In this embodiment, in the physiologically active composition of the present disclosure, the content of the leaf extract of the Saururus plant of the present disclosure may be about 10 - 99.5 wt%, for example, about 10 - 50 wt%, about 50 - 99.5 wt%, etc., but is not limited thereto.

[0093] The pharmaceutically acceptable carrier may include, but is not limited to, solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic agents and absorption delaying agents, etc., and those suitable for pharmaceutical administration. According to different dosing methods, pharmaceutical compositions can be prepared into dosage forms using common methods.

[0094] The pharmaceutically acceptable salts include, but are not limited to, salts containing inorganic cations, such as, by way of example, alkali metal salts such as sodium salts, potassium salts or amine salts, alkaline earth metal salts such as magnesium salts, calcium salts, salts containing divalent or tetravalent cations such as zinc salts, aluminum salts or zirconium salts. Also, organic salts such as, by way of example, dicyclohexylamine salts, methyl-D-glucamine, amino acid salts such as arginine, lysine, histidine, glutamine may be used.

[0095] The pharmaceutical compositions described in the present disclosure can be administered parenterally, orally, by inhalation spray, or by the method of an implanted reservoir. Parenteral administration includes rubbing on any part or the required part of the skin, subcutaneous, intracutaneous, intravenous, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional injection and infusion techniques.

[0096] In one embodiment, the pharmaceutical composition described in the present disclosure may be in an oral dosage form, and the oral dosage form may include, but is not limited to, tablets, granules, powders, pellet capsules, capsules, coated tablets, emulsions, aqueous solutions, aqueous suspensions, dispersions, drugs of the type that are dissolved in water or the like and taken orally.

[0097] In another embodiment, the pharmaceutical composition described in the present disclosure may be in a dosage form for topical use by rubbing, and the dosage form for topical use by rubbing may include, but is not limited to, ointments, emulsions, solutions, gels, mousses and the like.

[0098] In addition, in one embodiment, the bioactive composition of the present disclosure may be a health supplement composition, and, without limitation, may further contain food-acceptable additives. In this embodiment, in the bioactive composition of the present disclosure, the content of the leaf extract of the genus Banjiro plants of the present disclosure may be about 10 to 99.5 wt%, for example, about 10 to 50 wt%, about 50 to 99.5 wt%, etc., but is not limited thereto.

[0099] The food-acceptable additives may include, without limitation, preservatives, bactericides, antioxidants, bleaching agents, color developers, swelling agents, coloring agents, seasonings, sweeteners, thickeners (pastes), binders, emulsifiers, carriers, etc. or any combination thereof.

[0100] The health supplement composition described in the present disclosure may be in an oral form, and this oral form may include, but is not limited to, tablets, granules, powders, pellet capsules, capsules, coated tablets, emulsions, aqueous solutions, aqueous suspensions, dispersions, types that are dissolved in water and drunk, etc.

[0101] In addition, in one embodiment, the bioactive composition of the present disclosure may be a skin care composition, and, without limitation, may contain a pharmaceutically acceptable carrier. For the description of the pharmaceutically acceptable carrier, reference is made to the related description mentioned above and will not be repeated here. In this embodiment, in the bioactive composition of the present disclosure, the content of the leaf extract of the genus Banjiro plants of the present disclosure may be about 10 to 99.5 wt%, for example, about 10 to 50 wt%, about 50 to 99.5 wt%, etc., but is not limited thereto.

[0102] The skin care composition described in the present disclosure may be in a form for topical use by rubbing, and the form for topical use by rubbing may include, but is not limited to, ointments, emulsions, solutions, gels, mousses, etc.

[0103] In addition, the subjects to which the bioactive composition of the present disclosure is administered may include, but are not limited to, vertebrates. Further, the vertebrates may include, but are not limited to, fish, amphibians, reptiles, birds, or mammals. Examples of mammals may include, but are not limited to, humans, orangutans, monkeys, horses, donkeys, dogs, cats, rabbits, guinea pigs, rats, and mice. In one embodiment, the subject to which the bioactive composition of the present disclosure is administered may be a human.

Examples

[0104] A. Experimental method

[0105] A-1. Extraction method of leaves of Pandanus utilis

[0106] 1. Extraction method A (extraction only with a solvent) (1) From the branches and leaves of Pandanus utilis, leaves in good condition and not withered or rotten were selected, then they were separated from the branches, the petioles were removed, and the selected leaves were obtained. Then, the selected leaves were washed clean. (2) The washed leaves were baked at a constant temperature of 50 °C, and then the dried leaves were ground into powder. The size (particle diameter) of the powder was about 0.25 mm. (3) 1 g of the powder of Pandanus utilis leaves obtained above was mixed with 14 g of RO water to prepare a leaf powder suspension, and the pH value of the leaf powder suspension was adjusted to 5.0 with citric acid. (4) 6.5 g of ethanol was added to the above leaf powder suspension, and the extraction process was carried out by performing ultrasonic vibration at 60 °C for 1 hour to obtain an extract. (5) The obtained extract was first filtered through a filter paper with a pore size of 6 μm, and then filtered through a filter paper with a pore size of 1 μm to obtain a filtrate. (6) The obtained filtrate was centrifuged at 25 °C and 5000 rpm for 15 minutes. (7) After centrifugation, the supernatant was taken and freeze-dried into powder.

[0107] 2. Extraction method B (stirring for 1 hour + extraction with a solvent) (1) Select leaves from the branches and leaves of the banyan tree that are in good condition and not withered or rotten. Then, separate them from the branches, remove the petioles, and obtain the selected leaves. Next, wash these selected leaves clean. (2) Bake the washed leaves at a constant temperature of 50 °C, and then grind the dried leaves into powder. The size (particle size) of the powder was set to about 0.25 mm. (3) Mix 1 g of the powder of the banyan tree leaves obtained above with 14 g of RO water to make a leaf powder suspension, and adjust the pH value of the leaf powder suspension to 5.0 with citric acid. (4) Stir the above leaf powder suspension at 45 °C for 1 hour. (5) Then, add 6.5 g of ethanol to the above leaf powder suspension, and perform ultrasonic vibration at 60 °C for 1 hour to carry out the extraction process and obtain an extract. (6) First, filter the obtained extract with a filter paper having a pore size of 6 μm, and then filter it with a filter paper having a pore size of 1 μm to obtain a filtrate. (7) Centrifuge the obtained filtrate at 25 °C and 5000 rpm for 15 minutes. (8) After centrifugation, take the supernatant and freeze-dry it into powder.

[0108] 3. Extraction Method C (Use of Enzyme Formulation α) (1) Select leaves from the branches and leaves of the banyan tree that are in good condition and not withered or rotten. Then, separate them from the branches, remove the petioles, and obtain the selected leaves. Next, wash these selected leaves clean. (2) Bake the washed leaves at a constant temperature of 50 °C, and then grind the dried leaves into powder. The size (particle size) of the powder was set to about 0.25 mm. (3) Mix 1 g of the powder of the banyan tree leaves obtained above with 14 g of RO water to make a leaf powder suspension, and adjust the pH value of the leaf powder suspension to 5.0 with citric acid. (4) Add the enzyme preparation of formulation α (0.1 g of enzyme mixture (containing Viscozyme® L. arabinase, cellulase, β-glucanase, hemicellulase and xylanase)) and 0.1 g of pectinase to the above leaf powder suspension, stir at 45 °C for 1 hour to allow the enzymatic depolymerization reaction to proceed, and obtain an enzyme-treated product. (5) Add 6.5 g of ethanol to the above enzyme-treated product, perform ultrasonic vibration at 60 °C for 1 hour to carry out the extraction process, and obtain an extract. (6) First filter the obtained extract with a filter paper having a pore size of 6 μm, and then filter it with a filter paper having a pore size of 1 μm to obtain a filtrate. (7) Centrifuge the obtained filtrate at 25 °C and 5000 rpm for 15 minutes. (8) After centrifugation, take the supernatant and lyophilize it to make a powder.

[0109] 4. Extraction method D (use of enzyme formulation β) (1) Select leaves from the branches and leaves of Banjilou that are in good condition and not withered or rotten, then separate them from the branches, remove the petioles to obtain selected leaves. Then wash these selected leaves clean. (2) Bake the washed leaves at a constant temperature of 50 °C, and then grind the dried leaves into powder. The size (particle size) of the powder was about 0.25 mm. (3) Mix 1 g of the powder of Banjilou leaves obtained above with 14 g of RO water to make a leaf powder suspension, and adjust the pH value of the leaf powder suspension to 5.0 with citric acid. (4) Add the enzyme preparation of formulation β (0.1 g of tannase) to the above leaf powder suspension, stir at 45 °C for 1 hour to allow the enzymatic depolymerization reaction to proceed, and obtain an enzyme-treated product. (5) Add 6.5 g of ethanol to the above enzyme-treated product, perform ultrasonic vibration at 60 °C for 1 hour to carry out the extraction process, and obtain an extract. (6) First filter the obtained extract with a filter paper having a pore size of 6 μm, and then filter it with a filter paper having a pore size of 1 μm to obtain a filtrate. (7) The obtained filtrate was centrifuged at 25 °C and 5000 rpm for 15 minutes. (8) After centrifugation, the supernatant was taken and freeze-dried to obtain a powder.

[0110] 5. Extraction method E (use of enzyme formulation γ) (1) From the branches and leaves of the pandanus, leaves in good condition and not withered or rotten were selected. Then, they were separated from the branches, the petioles were removed to obtain the selected leaves. Next, the selected leaves were washed clean. (2) The washed leaves were baked at a constant temperature of 50 °C, and then the dried leaves were ground into powder. The size (particle size) of the powder was about 0.25 mm. (3) 1 g of the powder of pandanus leaves obtained above was mixed with 14 g of RO water to prepare a leaf powder suspension, and the pH value of the leaf powder suspension was adjusted to 5.0 with citric acid. (4) The enzyme preparation of formulation γ (0.1 g of enzyme mixture (including Viscozyme (registered trademark) L. arabinase, cellulase, β-glucanase, hemicellulase and xylanase), 0.1 g of pectinase and 0.1 g of tannase) was added to the above leaf powder suspension, and stirred at 45 °C for 1 hour to allow the enzyme depolymerization reaction to proceed, obtaining an enzyme-treated product. (5) 6.5 g of ethanol was added to the above enzyme-treated product, and the extraction process was carried out by performing ultrasonic vibration at 60 °C for 1 hour to obtain an extract. (6) The obtained extract was first filtered through a filter paper with a pore size of 6 μm, and then filtered through a filter paper with a pore size of 1 μm to obtain a filtrate. (7) The obtained filtrate was centrifuged at 25 °C and 5000 rpm for 15 minutes. (8) After centrifugation, the supernatant was taken and freeze-dried to obtain a powder.

[0111] A-2. Method for measuring the content of active ingredients

[0112] 1. Method for measuring the total polysaccharide content

[0113] The total polysaccharide content in the extract of *Banzhilou* leaves was measured by the 3,5-dinitrosalicylic acid (DNS) method.

[0114] When 3,5-dinitrosalicylic acid is heated with reducing sugar, it is reduced to a brown amino compound. Using a spectrophotometer, the absorbance value of this brown amino compound can be measured at a wavelength of 540 nm. Within a certain range, the color intensity of this brown amino compound is directly proportional to the reducing sugar concentration. If reducing sugar is present, the color changes from yellow to brown. Therefore, since the reducing sugar concentration in the sample can be measured using the colorimetric method, the total polysaccharide content in the sample can also be measured.

[0115] (1) Preparation of glucose calibration curve 1 mL of glucose solutions with different concentrations (0, 100, 200, 400, 800 μg / mL) was uniformly mixed with 1 mL of DN reagent respectively to prepare reaction solutions. These reaction solutions were heated at 100 °C for 10 minutes, cooled, and then placed in an ice box to lower the temperature. Then, the reaction solutions were added to 3 mL of distilled water, and their absorbance values were measured at a wavelength of 540 nm. A glucose concentration calibration curve was prepared by correlating each concentration of glucose solution with its absorbance value, and an equation corresponding to the calibration curve was obtained.

[0116] (2) Measurement of the total polysaccharide content in the extract of *Banzhilou* leaves The powder of the extract of *Banzhilou* leaves was prepared into a sample solution with a concentration of 10000 ppm with water.

[0117] 1 mL of the sample solution was uniformly mixed with 1 mL of DNS reagent to prepare reaction solutions respectively. These reaction solutions were heated at 100 °C for 10 minutes, cooled, and then placed in an ice box to lower the temperature. Then, the reaction solutions were added to 3 mL of distilled water, and their absorbance values were measured at a wavelength of 540 nm. The absorbance value of the sample solution was applied to the equation of the calibration curve to calculate the corresponding total polysaccharide content.

[0118] 2. Method for Measuring the Contents of Ellagic Acid, Gallic Acid, and Quercetin

[0119] (1) Preparation of Calibration Curve for the Substance to be Measured Standard substances (ellagic acid standard, gallic acid standard, or quercetin standard) of the substance to be measured at different concentrations (100 ppm, 50 ppm, 25 ppm, 12.5 ppm, 6.25 ppm) were prepared with ethanol.

[0120] Using a C18 chromatography column, the standard substances at the above different concentrations were measured by high performance liquid chromatography (HPLC). The conditions of high performance liquid chromatography were as follows. Detection wavelength: 270 nm Mobile phase A: 0.1% acetic acid + water Mobile phase B: 0.1% acetic acid + acetonitrile Flow rate: 1 mL / min Sample injection volume: 10 μL Analysis gradient: 0 - 5 minutes, mobile phase A 90%, mobile phase B 10%; 5 - 30 minutes, mobile phase A from 90% to 0%, mobile phase B from 10% to 100%; 30 - 35 minutes, mobile phase A 0%, mobile phase B 100%; 35 - 35.5 minutes, mobile phase A from 0% to 90%, mobile phase B from 100% to 10%; 35.5 - 40 minutes, mobile phase A 90%, mobile phase B 10%.

[0121] Based on the UV absorption spectrum at 270 nm, the integrated area of the characteristic peaks of the standard substances at different concentrations was calculated, a calibration curve of concentration and the integrated area of the characteristic peaks was prepared, and the corresponding calibration curve equation was obtained.

[0122] (2) Measurement of the Contents of Ellagic Acid, Gallic Acid, and Quercetin in the Extract of Banjiro Leaves The powder of the extract of Banjiro leaves was prepared into a sample solution with a concentration of 10000 ppm with ethanol.

[0123] For the sample solution, after obtaining its UV absorption spectrum at 270 nm by high performance liquid chromatography under the same conditions as above, the integrated area of its characteristic peak was calculated. The integrated area of the characteristic peak of the calculated sample solution was applied to the calibration curve equation to calculate the content of the corresponding substance to be measured.

[0124] B. Experimental Results

[0125] B-1. Obtaining Extracts of Banjirow Leaf

[0126] Extracts A to E of Banjirow leaf were obtained respectively by extraction methods A to E described in the "Measurement Method of Active Ingredient Content" in the above-mentioned "A. Experimental Method".

[0127] Thereafter, the contents of the active ingredients of extracts A to E of Banjirow leaf and commercially available Banjirow leaf extract were measured.

[0128] B-2. Measurement of the Contents of Active Ingredients of Extracts A to E of Banjirow Leaf and Commercially Available Banjirow Leaf Extract

[0129] 1. Measurement of the Total Polysaccharide Content of Extracts A to E and Commercially Available Banjirow Leaf Extract

[0130] According to the "1. Measurement Method of Total Polysaccharide Content" described in the "A-2. Extraction Method of Banjirow Leaf" in the above-mentioned "A. Experimental Method", the total polysaccharide contents of extracts A to E of Banjirow leaf and commercially available Banjirow leaf extract (Guava Leaf Extract Powder, Bizen Kasei Co., Ltd. (Japan)) were measured respectively.

[0131] The results are shown in Figure 1.

[0132] As shown in Fig. 1, compared with the extract A of banjirou leaves obtained by extraction method A that only extracts with a solvent, the extract B of banjirou leaves obtained by extraction method B that stirs for 1 hour and extracts with a solvent, the commercially available banjirou leaf extract, the extract C of banjirou leaves obtained by extraction method C that is treated with enzyme formulation α and extracts with a solvent, the extract D of banjirou leaves obtained by extraction method D that is treated with enzyme formulation β and extracts with a solvent, and the extract E of banjirou leaves obtained by extraction method E that is treated with enzyme formulation γ and extracts with a solvent all had a higher total polysaccharide content.

[0133] Compared with extraction method B that stirs for 1 hour and extracts with a solvent, extraction method E that is treated with enzyme formulation γ and extracts with a solvent was able to increase the total polysaccharide extraction amount by 137.8%. Also, compared with the commercially available banjirou leaf extract, extraction method E that is treated with enzyme formulation γ and extracts with a solvent was able to increase the total polysaccharide extraction amount by 147.2%.

[0134] 2. Measurement of ellagic acid content in extracts A to E and the commercially available banjirou leaf extract

[0135] According to the "2. Measurement method of ellagic acid, gallic acid, and quercetin content" described in "A-2. Extraction method of banjirou leaves" in the aforementioned "A. Experimental method", the ellagic acid contents of extracts A to E of banjirou leaves and the commercially available banjirou leaf extract (Guava Leaf Extract Powder, Bizen Kasei Co., Ltd. (Japan)) were measured respectively.

[0136] The results are shown in Fig. 2.

[0137] As shown in Fig. 2, compared with the extract B of banjirou leaves obtained by extraction method B of stirring for 1 hour and extracting with a solvent, the extract A of banjirou leaves obtained by extraction method A of only extracting with a solvent, the extract D of banjirou leaves obtained by extraction method D of treating with enzyme formulation β and extracting with a solvent, and the extract E of banjirou leaves obtained by extraction method E of treating with enzyme formulation γ and extracting with a solvent all had higher ellagic acid contents.

[0138] Compared with extraction method B of stirring for 1 hour and extracting with a solvent, extraction method D of treating with enzyme formulation β and extracting with a solvent was able to increase the ellagic acid extraction amount by 47.9%.

[0139] Also, compared with extraction method B of stirring for 1 hour and extracting with a solvent, extraction method E of treating with enzyme formulation γ and extracting with a solvent was able to increase the ellagic acid extraction amount by 32.7%.

[0140] 3. Measurement of Gallic Acid Contents in Extracts A to E and Commercially Available Banjirou Leaf Extracts

[0141] According to the "2. Measurement Method of Ellagic Acid, Gallic Acid, and Quercetin Contents" described in "A-2. Extraction Method of Banjirou Leaves" in the "A. Experimental Method" described above, the gallic acid contents of extracts A to E of banjirou leaves and a commercially available banjirou leaf extract (Guava Leaf Extract Powder, Bizen Kasei Co., Ltd. (Japan)) were measured respectively.

[0142] The results are shown in Fig. 3.

[0143] As shown in Fig. 3, the extract A of Psidium guajava leaves obtained by extraction method A which only extracts with a solvent, the extract B of Psidium guajava leaves obtained by extraction method B which stirs for 1 hour and extracts with a solvent, and compared with a commercially available Psidium guajava leaf extract, the extract D of Psidium guajava leaves obtained by extraction method D which is treated with enzyme formulation β and extracted with a solvent, and the extract E of Psidium guajava leaves obtained by extraction method E which is treated with enzyme formulation γ and extracted with a solvent all had higher gallic acid contents.

[0144] Compared with extraction method B which stirs for 1 hour and extracts with a solvent, extraction method D which is treated with enzyme formulation β and extracted with a solvent could increase the gallic acid extraction amount by 557.0%. Also, compared with a commercially available Psidium guajava leaf extract, extraction method D which is treated with enzyme formulation β and extracted with a solvent could similarly increase the gallic acid extraction amount by 557.0%.

[0145] Also, compared with extraction method B which stirs for 1 hour and extracts with a solvent, extraction method E which is treated with enzyme formulation γ and extracted with a solvent could increase the gallic acid extraction amount by 528.4%. Furthermore, compared with a commercially available Psidium guajava leaf extract, extraction method E which is treated with enzyme formulation γ and extracted with a solvent could similarly increase the gallic acid extraction amount by 528.4%.

[0146] 4. Measurement of quercetin contents in extracts A to E and a commercially available Psidium guajava leaf extract

[0147] According to the "2. Measurement method for contents of ellagic acid, gallic acid and quercetin" described in "A-2. Extraction method of Psidium guajava leaves" in the above-mentioned "A. Experimental method", the quercetin contents of extracts A to E of Psidium guajava leaves and a commercially available Psidium guajava leaf extract (Guava Leaf Extract Powder, Bizen Kasei Co., Ltd. (Japan)) were measured respectively.

[0148] The results are shown in Fig. 4.

[0149] As shown in FIG. 4, compared with extract A of Banjiro leaves obtained by extraction method A which only extracts with a solvent, and extract B of Banjiro leaves obtained by extraction method B which stirs for 1 hour and extracts with a solvent, extract C of Banjiro leaves obtained by extraction method C which treats with enzyme formulation α and extracts with a solvent, extract D of Banjiro leaves obtained by extraction method D which treats with enzyme formulation β and extracts with a solvent, and extract E of Banjiro leaves obtained by extraction method E which treats with enzyme formulation γ and extracts with a solvent all had higher quercetin contents.

[0150] Also shown in FIG. 4 is that compared with commercially available Banjiro leaf extract, extract C of Banjiro leaves obtained by extraction method C which treats with enzyme formulation α and extracts with a solvent, and extract E of Banjiro leaves obtained by extraction method E which treats with enzyme formulation γ and extracts with a solvent both had higher quercetin contents.

[0151] Also, compared with extraction method B which stirs for 1 hour and extracts with a solvent, extraction method E which treats with enzyme formulation γ and extracts with a solvent was able to increase the quercetin extraction amount by 411.9%. Furthermore, compared with commercially available Banjiro leaf extract, extraction method E which treats with enzyme formulation γ and extracts with a solvent was able to increase the quercetin extraction amount by 79.2%.

[0152] As can be seen from the above experimental results, by using the above combination of enzyme formulation and solvent extraction, the extraction amounts of active ingredients in Banjiro leaves, such as polysaccharides and polyphenols, can be more effectively increased.

[0153] Although the present disclosure has been disclosed as above according to preferred embodiments, these do not limit the present disclosure. Those skilled in the art can naturally make some changes and variations without departing from the spirit and scope of the present disclosure. Therefore, the protection scope of the present disclosure should be based on that defined by the appended claims.

Claims

1. A method for extracting leaves of plants of the genus Psidium, comprising: (a) preparing crushed pieces or powder of leaves of plants of the genus Psidium; (b) mixing the crushed pieces or powder of leaves of plants of the genus Psidium with water at a weight ratio of 1:7 to 21 to form a leaf crushed piece mixture or a leaf powder suspension; (c) adding an enzyme preparation to the leaf crushed piece mixture or the leaf powder suspension to form a first mixture, and performing an enzyme treatment step for 0.5 to 24 hours to obtain an enzyme-treated product, wherein the weight ratio of the crushed pieces or powder of leaves of plants of the genus Psidium to the enzyme preparation is 1:0.01 to 1, and the formulation of the enzyme preparation includes one of the formulations shown in the following (i) to (iv); (i) Formulation (I) containing an enzyme mixture, wherein the enzyme mixture includes arabinase, cellulase, β-glucanase, hemicellulase, and xylanase; (ii) Formulation (II) containing the enzyme mixture and pectinase; (iii) Formulation (III) containing tannase; (iv) Formulation (IV) containing the enzyme mixture, the pectinase, and the tannase; (d) mixing the enzyme-treated product with an alcohol solvent at a weight ratio of 1:0.1 to 2.5 to form a second mixture, and performing an extraction step to obtain an extract as an extract of leaves of plants of the genus Psidium, wherein the extract contains at least one active ingredient. A method for extracting leaves of plants of the genus Psidium, comprising the above steps.

2. The method for extracting leaves of plants of the genus Psidium according to claim 1, wherein the plants of the genus Psidium include Psidium guajava, Psidium littorale, or Psidium acutangulum.

3. The method for extracting leaves of plants of the genus Psidium according to claim 1, wherein the plants of the genus Psidium are Psidium guajava.

4. The at least one active ingredient includes at least one of the following components: Polysaccharides and polyphenols The method for extracting leaves of plants of the genus Psidium according to claim 1, wherein the method includes at least one of the above components.

5. The polyphenols include the following polyphenols: Phenolic acid, flavonoid, stilbenoid, and lignan The method for extracting leaves of the genus Banjirow, according to claim 4, wherein at least one of them is included.

6. Among the phenolic acids, the following phenols: Ellagic acid, gallic acid, and quinic acid At least one of which is included, and Among the flavonoids, the following flavonoids: Quercetin and kaempferol The method for extracting leaves of the genus Banjirow, according to claim 5, wherein at least one of them is included.

7. In the step (b), the pH value of the leaf crushed fragment mixture or the leaf powder suspension is adjusted to 4.5 to 5.

5. The method for extracting leaves of the genus Banjirow, according to claim 1.

8. In the step (c), the enzyme treatment step is carried out at 35 to 55°C. The method for extracting leaves of the genus Banjirow, according to claim 1.

9. In the step (c), the weight ratio of the crushed fragments or powder of the leaves of the genus Banjirow to the enzyme preparation is 1:0.1 to 0.

3. The method for extracting leaves of the genus Banjirow, according to claim 1.

10. In the formulation (II), the weight ratio of the enzyme mixture to the pectinase is 1:0.5 to 2. The method for extracting leaves of the genus Banjirow, according to claim 1.

11. In the formulation (IV), the weight ratio of the enzyme mixture, the pectinase, and the tannase is 1:0.5 to 2:0.5 to 2. The method for extracting leaves of the genus Banjirow, according to claim 1.

12. In the step (d), in the second mixture, the final concentration of the alcohol solvent is 20 to 50%. The method for extracting leaves of the genus Banjirow, according to claim 1.

13. In the step (d), the alcohol solvent includes methanol, ethanol, ethylene glycol, propanol, or isopropanol. The method for extracting leaves of the genus Banjirow, according to claim 1.

14. In the step (d), the extraction step is carried out at 50 to 70°C for 0.5 to 24 hours. The method for extracting leaves of the genus Banjirow, according to claim 1.

15. A leaf extract of a Pandanus plant, which contains at least one physiologically active ingredient and is obtained by the method for extracting leaves of a Pandanus plant according to any one of claims 1 to 14.

16. A physiologically active composition containing the leaf extract of a Pandanus plant according to claim 15.

17. A pharmaceutical composition, which is the physiologically active composition according to claim 16 and further contains a pharmaceutically acceptable carrier or salt.

18. A health supplement food composition, which is the physiologically active composition according to claim 16 and further contains a food-acceptable additive.

19. A skin care composition, which is the physiologically active composition according to claim 16 and further contains a pharmaceutically acceptable carrier.

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