Antioxidant, method for producing antioxidant, α-glucosidase inhibitor, and method for producing α-glucosidase inhibitor

Abstract

To provide an antioxidant activator having excellent functionality of an extract obtained from olive, a method for producing the activator, α-glucosidase inhibition activator, and a method for producing the activator.SOLUTION: An antioxidant activator contains a mycelium-processed product that is obtained by processing an extract obtained from branches and / or leaves of olive by a mycelium. By containing the mycelium-processed product in which the extract obtained from olive is processed by a mycelium, excellent antioxidant activity can be exerted, so that effective prevention of cancer, lifestyle-related diseases, infectious diseases, dementia, or the like is expected. In addition, unused biomass resources can be effectively utilized.SELECTED DRAWING: Figure 1

Description

Technical Field

[0001] The present invention relates to an antioxidant, a method for producing an antioxidant, an α-glucosidase inhibitor, and a method for producing an α-glucosidase inhibitor. More specifically, it relates to an antioxidant using olive branches and leaves, a method for producing an antioxidant, an α-glucosidase inhibitor, and a method for producing an α-glucosidase inhibitor.

Background Art

[0002] In recent years, the demand for health foods (including pharmaceuticals) aimed at maintaining and promoting health, mainly among middle-aged and elderly people, has been increasing. Since olive fruits contain various polyphenols such as oleuropein, they are used in a wide variety of functional foods. For example, Patent Document 1 discloses an α-glucosidase inhibitor using a component extracted from the aqueous solution obtained by pressing olive fruits, and Patent Document 2 discloses an amylase inhibitor using olive leaves and its extract.

Prior Art Documents

Patent Documents

[0003]

Patent Document 1

Patent Document 2

Disclosure of the Invention

Problems to be Solved by the Invention

[0004] However, the technologies of Patent Documents 1 and 2 are merely technologies that use the components contained in olives as they are without any chemical modification. Therefore, due to the recent increasing trend towards health consciousness, the development of health foods that can more efficiently ingest the functions of olives that improve human health functions is expected.

[0005] In view of the above circumstances, an object of the present invention is to provide an antioxidant having improved functionality of an extract obtained from olives, a method for producing the same, an α-glucosidase inhibitor, and a method for producing the same.

Means for Solving the Problems

[0006] As a result of intensive studies to solve the above problems, the present inventors have found that the functionality of an extract obtained from olives can be improved by treating the extract with a mycelium, and have completed the present invention.

[0007] The antioxidant of the present invention is characterized by containing a mycelium-treated product obtained by treating an extract obtained from olive branches and / or leaves with a mycelium. The method for producing the antioxidant of the present invention comprises a first step of separating into a distillate and a solid content by vacuum distillation while irradiating olive branches and / or leaves with microwaves, a second step of extracting the obtained solid content with water or hot water to obtain an extract, and a third step of treating the obtained extract with a mycelium, which are carried out in this order. The α-glucosidase inhibitor of the present invention is characterized by containing a mycelium-treated product obtained by treating an extract obtained from olive branches and / or leaves with a mycelium. The method for producing the α-glucosidase inhibitor of the present invention comprises a first step of separating into a distillate and a solid content by vacuum distillation while irradiating olive branches and / or leaves with microwaves, a second step of extracting the obtained solid content with water or hot water to obtain an extract, and a third step of treating the obtained extract with a mycelium, which are carried out in this order.

[0008] According to the present invention, by treating an extract obtained from olives with a mycelium, it is possible to provide an antioxidant, an α-glucosidase inhibitor, and methods for producing the same, which are expected to efficiently prevent cancer, lifestyle-related diseases, infectious diseases, dementia, and the like.

Brief Description of the Drawings

[0009]

Figure 1

Figure 2

Figure 3

Figure 4

Mode for Carrying Out the Invention

[0010] Embodiments of the present invention will be described based on the drawings. The antioxidant of the present embodiment contains a mycelium-treated product obtained by treating an extract obtained from olive branches and leaves with mycelium, and thus efficient prevention of cancer, infectious diseases, dementia, etc. is expected. The α-glucosidase inhibitor of the present embodiment contains a mycelium-treated product obtained by treating an extract obtained from olive branches and leaves with mycelium, and thus prevention of lifestyle-related diseases and control of blood glucose levels are expected.

[0011] In the specification, an olive is a plant belonging to the genus Olea of the family Oleaceae, and its type is not particularly limited. The parts of the olive to be targeted are branches and leaves, and these can be used alone or in combination. Hereinafter, when simply referred to as an olive, it means the above-mentioned target parts.

[0012] Also, the cultivation state of the olive to be treated is not particularly limited. For example, if olive branches and leaves that are planned to be discarded during the harvesting process of olive fruits are used, the advantage of effectively utilizing unused biomass resources can be obtained.

[0013] The mycelium-treated product is obtained by treating an extract obtained from olives with the mycelium of mushrooms. The mushrooms used for preparing the mycelium-treated product are not particularly limited. For example, shiitake mushrooms, enoki mushrooms, nameko mushrooms, oyster mushrooms, beech mushrooms, eryngii mushrooms, yamabushitake mushrooms, etc. can be mentioned.

[0014] The antioxidant agent of this embodiment (hereinafter simply referred to as the antioxidant agent) contains a mycelium-treated product obtained by treating an extract obtained from olives with mycelium. And by containing this mycelium-treated product, the antioxidant agent can exhibit excellent antioxidant activity. Therefore, by ingesting this antioxidant agent, it is expected to suppress the action of reactive oxygen generated in the body. Specifically, by ingesting the antioxidant agent, it is expected to be effective in preventing aging phenomena, cancer, infectious diseases, dementia, etc. caused by the influence of reactive oxygen.

[0015] The α-glucosidase inhibitor of this embodiment (hereinafter simply referred to as the α-glucosidase inhibitor) contains a mycelium-treated product obtained by treating an extract obtained from olives with mycelium, similar to the antioxidant agent. And by containing this mycelium-treated product, the α-glucosidase inhibitor is expected to suppress and regulate the digestion and absorption of carbohydrates by exerting α-glucosidase inhibitory activity in the body. Specifically, by ingesting the α-glucosidase inhibitor, it is expected to be effective in improving the constitution that is prone to taking in carbohydrates, preventing lifestyle-related diseases such as excessive carbohydrate intake, and controlling postprandial hyperglycemia.

[0016] Next, the manufacturing methods of the antioxidant agent and the α-glucosidase inhibitor will be described. Note that since the manufacturing methods of the antioxidant agent and the α-glucosidase inhibitor are the same, the manufacturing method of the antioxidant agent of this embodiment, which is the manufacturing method of the antioxidant agent (hereinafter simply referred to as the manufacturing method of the antioxidant agent), will be described as a representative.

[0017] A method for producing an antioxidant can produce the antioxidant by including a mycelium-treated product obtained by treating an extract obtained from olives with a mycelium.

[0018] First, the extract to be treated with the mycelium in the method for producing an antioxidant will be described. The extract to be treated with the mycelium is obtained from olives. Specifically, this extract contains components resulting from the solid content obtained after separating at least a part of the essential oil and / or aqueous liquid from the olives. For example, it includes solid forms containing the residue (solid content) itself, which is the residue obtained by performing treatments such as pressing or distillation, liquid forms in which this solid content is dispersed, and those containing a liquid obtained by further extracting this solid content, etc., can be cited as extracts.

[0019] The above-described method for preparing the solid content is not particularly limited as long as at least a part of the essential oil and / or aqueous liquid can be separated from the olives. For example, the solid content can be prepared using the above-described pressing machine, distillation apparatus, etc.

[0020] When preparing the solid content using a pressing machine, distillation apparatus, etc., it can be processed and supplied in a size that can be accommodated in the housing part of the apparatus, etc. Although olive leaves, etc. can be supplied as they are, from the viewpoints of processing time and processing efficiency, it is preferable to supply them in a reduced state. Known methods can be used for the method of reducing the size when supplying olives to the apparatus, etc. For example, by performing treatments such as cutting or pulverizing, the olives can be supplied to the apparatus as a pulverized product such as chips or powder.

[0021] When using a pressing machine, the above-mentioned essential oil and / or aqueous liquid can be obtained as the pressed juice, and the solid content can be obtained as the residue. In addition, when using a distillation apparatus, the above essential oil and / or aqueous liquid can be obtained as a distillate, and solids can be obtained as a residue which is the residue. That is, when using a distillation apparatus, the solids are the residue remaining after at least a part of the essential oil and / or aqueous liquid has been separated as a distillate from the olives supplied to the distillation apparatus by distilling the olives.

[0022] Also, after irradiating the olives with microwaves described later, solids may be obtained using a press. In this case, since the cell walls constituting the olives are in a state where they are easily broken by microwaves, the desired solids can be easily obtained. Note that by irradiating the olives with microwaves, a microwave irradiation-treated product in which a part of the essential oil and aqueous liquid contained in the olives has flowed out can be obtained. Since the cell walls constituting the olives are in a state where they are easily broken by microwaves in this microwave irradiation-treated product, it is also possible to use it as solids in this state.

[0023] In the treatment using a distillation apparatus, solids can be obtained by treatments such as atmospheric distillation, vacuum distillation, and steam distillation. Also, the heating method when using a distillation apparatus is not particularly limited, and methods such as irradiating with a heater, hot water, steam, and microwaves can be adopted.

[0024] For example, if the method of irradiating with microwaves is adopted, it is preferable because the components contained in the olives can be made into a state in which the mycelium can easily process them. Specifically, when microwaves are irradiated onto the olives, the moisture contained in the olives is heated and released as water vapor. At this time, essential oils and water-soluble compounds (including volatile components) contained in the olives are obtained as a distillate. On the other hand, the cell walls constituting the olives are in a state where they are easily broken by microwaves. Therefore, components that are difficult to extract by a general distillation method can be included in the residue (that is, solids) in a state where they are easily extracted. Therefore, if an extract containing solid matter prepared by adopting a method of irradiating microwaves is used, it becomes easier to process the components contained in olives by the mycelium. In particular, when preparing an extract by subjecting the solid matter obtained by the treatment using a distillation apparatus to the extraction treatment described later, an advantage is obtained in that it becomes easier to extract the components contained in the solid matter.

[0025] As the distillation apparatus for irradiating microwaves, a known one can be adopted. For example, a microwave vacuum distillation apparatus 1 as shown in FIG. 1 can be used. This microwave vacuum distillation apparatus 1 includes a distillation tank 2 corresponding to a housing portion for housing olives as a raw material, a microwave heating apparatus 3 for irradiating microwaves, an air flow inlet pipe 4 for supplying an inert gas such as air or nitrogen gas to the distillation tank 2, a distillate outflow pipe 5 for guiding the distilled distillate, a cooling apparatus 6 for cooling the distillate outflow pipe 5, a heating control apparatus 7 for controlling the temperature of the distillation tank 2, and a pressure control apparatus 10 for controlling the pressure inside the distillation tank 2. The pressure control apparatus 10 includes a vacuum pump 8 and a pressure control valve 9. Due to the configuration as described above, in this microwave vacuum distillation apparatus 1, the pressure inside the distillation tank 2 and the like are adjusted via the heating control apparatus 7 and the pressure control apparatus 10, respectively.

[0026] In addition, as the distillation conditions when using a distillation apparatus, known conditions can be adopted. The conditions vary depending on the internal volume, its moisture content, the intensity of microwaves, etc. For example, the pressure inside the housing portion of the apparatus can be adjusted to be under a reduced pressure of 10 to 90 kPa, and the treatment time can be adjusted to be 0.2 to 8 hours.

[0027] In addition, the mixture of essential oil and aqueous liquid obtained as the distillate can be separated into an oily fraction and a water-soluble fraction by performing treatments such as standing, redistillation, and liquid-liquid separation treatment. Among the separated fractions, the oily fraction corresponds to the essential oil described above, and the water-soluble fraction corresponds to the aqueous liquid described above. These fractions can be used for respective desired applications by separation.

[0028] As the extract, as described above, solid-shaped ones containing the solid content itself prepared by the above method, liquid-shaped ones in which the prepared solid content is dispersed in a solution, ones containing a liquid obtained by further subjecting the prepared solid content to an extraction treatment, etc. can be adopted. That is, any extract is prepared so as to contain the components contained in the solid content. First, the first extract can be easily prepared by aliquoting a predetermined amount of the prepared solid content. The second extract is prepared by impregnating the prepared solid content in a liquid such as water that does not affect the mycelium of the mushroom so that the components contained in the solid content are in a state where they can be easily extracted. And the last extract is prepared to contain a liquid obtained by extracting only the components contained in the prepared solid content.

[0029] The method for preparing the last extract is not particularly limited as long as it is a method capable of extracting the components contained in the solid content. For example, general extraction methods using water, hot water, organic solvents, etc. can be adopted. When performing an extraction treatment with water or hot water, after performing an extraction treatment with each liquid for a predetermined time, the supernatant obtained using a centrifuge can be adopted as the extract. Also, when performing an extraction treatment with an organic solvent, it is necessary to perform a treatment that does not affect the mycelium. For example, after removing the organic solvent from the liquid after extraction, it can be made into an extract by replacing it with a solvent that does not affect the mycelium such as water.

[0030] Next, inoculate the desired mushroom spawn into the extract prepared as described above. After inoculation, the mycelium-treated product can be prepared by culturing the mycelium for a predetermined time. This mycelium-treated product is a mycelium-treated product obtained by treating an extract obtained from olives contained in an antioxidant and an α-glucosidase inhibitor with the mycelium.

[0031] In summary, the method for producing an antioxidant is a production method that sequentially performs a step of preparing an extract obtained from olives and a step of treating the prepared extract with a mycelium. The former step of preparing an extract obtained from olives includes a step of preparing a solid content obtained from olives. This step includes a step of adjusting the solid content by microwave irradiation treatment, a step of preparing the solid content by a pressing method, or a step of preparing the solid content by a distillation method. And in the step of preparing the solid content by the distillation method, a method of performing vacuum distillation while irradiating with microwaves is included. Also, when further extracting the solid content, it can include a step of further performing an extraction treatment on the prepared solid content using water, hot water, or the like.

[0032] Note that the method of preparing a solid content from olives by vacuum distillation while irradiating with the above microwaves corresponds to the first step in the claims. And the step of further performing an extraction treatment on the prepared solid content using water, hot water, or the like corresponds to the second step in the claims.

[0033] By using the production method as described above, an antioxidant exhibiting excellent antioxidant activity can be produced. Also, by using a similar production method, an α-glucosidase inhibitor exhibiting excellent α-glucosidase inhibitory activity can be produced.

Examples

[0034] Next, the present invention will be described in more detail with reference to examples. Note that these examples show an example of this embodiment, and the present invention is not limited in any way by the following examples.

[0035] The pulverized product obtained by pulverizing the pruned branches and leaves of olives with a pulverizer was put into the distillation tank of the microwave vacuum distillation apparatus shown in FIG. 1, and distillation treatment was performed under the condition that the inside of the distillation tank was set to the following conditions. Pressure inside the distillation tank: Vacuum condition of 0.25 KPa Distillation time: 30 minutes

[0036] Distillation was performed using the microwave vacuum distillation apparatus shown in Fig. 1 to obtain a residue (solid content) and a distillate.

[0037] The obtained residue was taken out of the distillation tank, a predetermined amount was collected, and extraction treatment was performed using water to prepare a water extract. Also, a predetermined amount was collected from the residue, and extraction treatment was performed using hot water to prepare a hot water extract.

[0038] Note that the prepared water extract and hot water extract correspond to the extract obtained from olives as referred to in this embodiment.

[0039] (Preparation of water extract) The preparation of the water extract was carried out as follows. 1. 50 ml of ultrapure water was added to 1 g of olive branches and leaves, and it was allowed to stand at 4°C for 24 hours. 2. Centrifugation (8000 rpm, 4°C, 30 minutes) was performed to collect the supernatant. 3. Centrifugation (8000 rpm, 4°C, 15 minutes) was performed again, and the collected supernatant was used as the water extract.

[0040] (Preparation of hot water extract) 1. 1 g of olive branches and leaves was added to 50 ml of boiling ultrapure water, and it was allowed to stand at 100°C for 20 minutes. 2. After cooling to room temperature, centrifugation (8000 rpm, 4°C, 30 minutes) was performed to collect the supernatant. 3. Centrifugation (8000 rpm, 4°C, 15 minutes) was performed again, and the collected supernatant was used as the hot water extract.

[0041] (Mycelium treatment) The following mycelium treatment was performed on the prepared water extract and hot water extract. Enoki mushroom was used as the type of mushroom for the mycelium. The mushroom inoculum was added to each extract, and it was allowed to stand at 25°C in the dark for 24 hours to prepare a mycelium-treated liquid.

[0042] Note that this mycelium treatment solution corresponds to a mycelium-treated product obtained by treating an extract obtained from olives as referred to in the present embodiment with mycelium.

[0043] Solids were filtered off from the prepared mycelium treatment solution to prepare a measurement sample for measuring antioxidant activity and α-glucosidase inhibitory activity.

[0044] (Analysis of antioxidant activity) Antioxidant activity was analyzed using the measurement sample. Antioxidant activity was evaluated in two ways: superoxide radical scavenging activity and DPPH radical scavenging activity.

[0045] (Superoxide radical scavenging activity) Measurement was performed by a luminescence method using a commercially available kit (ATTO Corporation, AB-2970 CLETA-S kit). The luminescence amount was measured using ATTO Corporation's luminescence measuring device AB-2270 luminometer.

[0046] (DPPH radical scavenging activity) 20 μl of a sample or extraction solvent (negative control) was added to 180 μl of a DPPH solution (1.0 mg of DPPH, 5.0 ml of 100% ethanol, 5.0 ml of 0.1 M MES buffer (pH 6.0)), and the mixture was reacted at room temperature in the dark for 30 minutes. After the reaction, the absorbance at 517 nm was measured using a microplate reader (manufactured by Corona Electric Co., Ltd., SH-1200 Lab).

[0047] (Analysis of α-glucosidase inhibitory activity) α-Glucosidase inhibitory activity was analyzed using the measurement sample. α-Glucosidase inhibitory activity was evaluated using α-glucosidase (manufactured by Sigma-Aldrich, product number; I1630). A predetermined amount was aliquoted from the sample for measurement, mixed with α-glucosidase, and then allowed to stand at room temperature for 10 minutes. Thereafter, an aqueous solution of p-nitrophenyl α-D-glucopyranoside was added to the adjusted mixture, and the reaction was carried out with stirring at 37 °C for 30 minutes. Then, an aqueous Tris solution was added, and the absorbance at 400 nm was measured using a microplate reader (SH-1200 Lab, manufactured by Corona Electric Co., Ltd.).

[0048] The α-glucosidase inhibitory activity was expressed as the α-glucosidase inhibitory activity rate (%). In the figure, it was simply shown as "inhibition rate (%)". The α-glucosidase inhibitory activity rate (%) was determined by dividing the value obtained by subtracting the blank value from each analytical value by the blank value.

[0049] The evaluation results of the antioxidant activity are shown in Fig. 2. In the superoxide radical scavenging activity of Fig. 2(A), for the water extract (olive branch and leaf water extraction system in the figure), the value before treatment (left graph) was 15.7% and the value after treatment (right graph) was 44.5%. Also, for the hot water extract (olive branch and leaf hot water extraction system in the figure), the value before treatment (left graph) was 30.8% and the value after treatment (right graph) was 58.8%. In the DPPH radical scavenging activity of Fig. 2(B), for the water extract (olive branch and leaf water extraction system in the figure), the value before treatment (left graph) was 23.1% and the value after treatment (right graph) was 35.1%. Also, for the hot water extract (olive branch and leaf hot water extraction system in the figure), the value before treatment (left graph) was 30.8% and the value after treatment (right graph) was 50.8%. As shown in Fig. 2, before (water extract or hot water extract) and after treating the mycelium (mycelium-treated solution), the value after treating the mycelium was improved by about 2-fold in activity compared to before treatment. Also, it was confirmed that the hot water extract had higher activity than the water extract.

[0050] The evaluation results of the α-glucosidase inhibitory activity (α-glucosidase inhibitory activity rate (%)) are shown in Fig. 3. In the α-glucosidase inhibitory activity shown in Fig. 3, in the water extract (olive branch and leaf water extraction system in the figure), the value before treatment (left graph) was 57.1%, and the value after treatment (right graph) was 68.2%. Also, in the hot water extract (olive branch and leaf hot water extraction system in the figure), the value before treatment (left graph) was 48.6%, and the value after treatment (right graph) was 90.5%. As shown in Fig. 3, similar to the evaluation of antioxidant activity, before and after treating the mycelium, an improvement in activity was confirmed for the value after mycelium treatment compared to before treatment. Also, it was confirmed that the hot water extract was more effective in this regard.

[0051] (HPLC analysis of the mycelium treatment solution) The mycelium treatment solution (using the hot water extract) was analyzed using HPLC (manufactured by Shimadzu Corporation). In the experiment, the mycelium treatment solution of Flammulina velutipes was used.

[0052] The measurement conditions were as follows. Column used: Shim-pack CLC-ODS 250×4.6mm (manufactured by Shimadzu Corporation) Mobile phase: Solution A (2% acetic acid), Solution B (100% methanol) Flow rate: 0.5 ml / min Measurement wavelength: 280 nm

[0053] The measurement results are shown in Fig. 4. Oleuropein, which is known to be contained in olive leaves before treatment, was not detected in the mycelium treatment solution after mycelium treatment. Also, peaks that were not seen before treatment were confirmed in the mycelium treatment solution after mycelium treatment. This suggests that oleuropein was denatured by the enzyme of the mycelium, and components different from oleuropein were produced. It is conceivable that these different components activated various health functions more than oleuropein. Note that the color of the liquid changed before (aqueous extract or hot water extract) and after (mycelium treatment solution) treating the mycelium. By treating the mycelium, the liquid turned brown. The browning indicates that polyphenols polymerized, and although it is unclear whether it is a substance detected by HPLC, it is conceivable that it contributed to the improvement of functionality.

Industrial Applicability

[0054] The antioxidant and α-glucosidase inhibitor of the present invention are suitable for preventing cancer, lifestyle-related diseases, infectious diseases, dementia, and the like. Further, the method for producing the antioxidant of the present invention and the method for producing the α-glucosidase inhibitor are each suitable for producing the antioxidant and α-glucosidase inhibitor of the present invention.

Explanation of Symbols

[0055] 1 Microwave vacuum distillation apparatus 2 Distillation tank 3 Microwave heating apparatus 4 Airflow inlet pipe 5 Distillate outflow pipe 6 Cooling apparatus 7 Heating control apparatus 8 Vacuum pump 9 Pressure control valve 10 Pressure control apparatus

Claims

1. It contains a mycelium-treated product obtained by adding a mycelium of the genus Flammulina to an extract obtained from olive branches and / or leaves, wherein the extract is a liquid, and is obtained by subjecting a residue obtained by separating at least a part of essential oil and / or an aqueous liquid by vacuum distillation while irradiating olive branches and / or leaves with microwaves to extraction treatment with water or hot water. An antioxidant characterized by the above.

2. A first step of separating into a distillate and a residue by vacuum distillation while irradiating olive branches and / or leaves with microwaves, a second step of extracting the obtained residue with water or hot water to obtain a liquid extract, and a third step of adding a mycelium of the genus Flammulina to the obtained extract are performed in order. A method for producing an antioxidant characterized by the above.

3. It contains a mycelium-treated product obtained by adding a mycelium of the genus Flammulina to a liquid extract obtained from olive branches and / or leaves, wherein the extract is obtained by subjecting a residue obtained by separating at least a part of essential oil and / or an aqueous liquid by vacuum distillation while irradiating olive branches and / or leaves with microwaves to extraction treatment with water or hot water. An α-glucosidase inhibitor characterized by the above.

4. A first step of separating into a distillate and a residue by vacuum distillation while irradiating olive branches and / or leaves with microwaves, a second step of extracting the obtained residue with water or hot water to obtain a liquid extract, and a third step of adding a mycelium of the genus Flammulina to the obtained extract are performed in order. A method for producing an α-glucosidase inhibitor characterized by the above.

Images