CN110167572B - Black raspberry extract, method for producing same, cataract prevention or progression inhibitor, and food and drink - Google Patents
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Abstract
The black raspberry extract of the invention induces the expression of alpha-crystal protein.
Description
Technical Field
The present invention relates to a black raspberry extract, a cataract-preventing or-developing inhibitor and a food or drink containing the black raspberry extract, a method for producing the black raspberry extract, and a method for preventing cataract or a method for suppressing the development thereof using the black raspberry extract.
Background
Oxygen is indispensable for most organisms, and several percent of oxygen taken into the body becomes active oxygen for defense against bacterial infection and the like. However, in order to maintain homeostasis, the residual active oxygen needs to be removed by antioxidase in the living body and exogenous antioxidant substances in food.
In addition, when an excessive amount of active oxygen is generated by ultraviolet rays, smoking, exhaust gas, air pollution, stress, or the like, oxidative damage is given to living tissues (see, for example, non-patent document 1). As a result, it can be said that 90% or more of diseases cause a causal relationship with the excessive production of active oxygen, such as cancer including lifestyle diseases and senile diseases, and dementia (see, for example, non-patent document 2).
Therefore, attention has been paid to activation of Superoxide dismutase (SOD) in vivo and antioxidant substances in foods, and a method for measuring antioxidant activity (ORAC; Oxygen Radical Absorbance Capacity) developed by the United States Department of Agriculture (USDA) has been established and is becoming widespread (see, for example, non-patent document 3). Further, there are 12 varieties of berry-like foods in the best 100 varieties of foods having high ORAC values, i.e., foods having high antioxidant ability (see, for example, non-patent document 4).
As an example of the senile disease, cataract may be mentioned. Cataract is caused by clouding of the lens, but it is not clear why the transparency of the lens is lost and clouding occurs. Maintenance of lens clarity is associated with the ordered structure of lens constituent proteins. When a Crystallin (Crystallin) as a constituent protein thereof is aggregated, clouding occurs to reduce the lens function of the lens, and the clouding affects the visual field and the eyesight (see, for example, non-patent document 5). In addition, among the crystallins, α -crystallins have been found to have a function of inhibiting the aggregation-binding of crystallins.
It can be said that the lens proteins of the eye are not substitutable for the whole life, and it is useful for the lens to supply the α -crystallin by gene expression again so as not to cause turbidity due to insolubilization.
Documents of the prior art
Non-patent document
Non-patent document 1: oberdanner, C.ROS and anticipatory Systems in Apoptosis.VDM Verlag,2008.ISBN 978-
Non-patent document 2: slave JL et al, health benefits of fruits and vegetables. adv nutr.2012.1; 3(4): 506-516
Non-patent document 3: cao G et al, oxy-radial affinity activity for antioxidants, free radial biol. med. 1993.14: 303-311
Non-patent document 4: "optimal 100 list of foods with high ORAC value (high antioxidant effect)" [ online (online) ] (search of 6 months and 30 days 2017) http: // www.anteaoxidant.com/orac # top100.html
Non-patent document 5: hipkiss, Accumulation of altered proteins and agents: causes and effects Experimental Geronology 41(2006)464-
Disclosure of Invention
Problems to be solved by the invention
The purpose of the present invention is to provide a berry extract having an action of preventing or inhibiting the progression of cataract, a cataract preventing or progression inhibitor and food and drink containing the berry extract, and a method for producing the berry extract. Another object of the present invention is to provide a method for preventing cataract or a method for inhibiting the progression thereof, using a berry extract.
The present inventors have focused on a berry food which is a food having an antioxidant effect, and have studied the correlation between the berry food and the effect of preventing or inhibiting the progression of cataract which is an senile disease, and as a result, have found that an extract of black raspberry achieves the above-mentioned object, and have completed the present invention.
Means for solving the problems
That is, according to the present invention, there can be provided:
(1) a black raspberry extract that induces the expression of alpha-crystallin;
(2) a cataract preventing or progressing inhibitor, which contains the black raspberry extract of (1);
(3) a food or drink comprising the black raspberry extract of (1);
(4) a method for preparing a black raspberry extract according to (1), comprising: freezing the fruit of the black raspberry; a thawing step of thawing the fruit of the black raspberry; a stirring step of adding a solvent to the fruit of the black raspberry thawed in the thawing step and stirring the mixture; and an extraction step of extracting the fruit of the black raspberry stirred by the stirring step by standing the fruit with a solvent;
(5) a method for preventing cataract or a method for inhibiting the progression thereof, wherein the black raspberry extract of (1) is used.
Effects of the invention
The present invention can provide a berry extract having an effect of preventing or inhibiting the progression of cataract, a cataract-preventing or progression-inhibiting agent and food and drink containing the berry extract, and a method for producing the berry extract. Further, the present invention can provide a method for preventing cataract or a method for inhibiting the progression thereof using the extract of berries.
Drawings
FIG. 1 is a graph showing induction of α -crystallin expression in a berry-like sample and Blabina.
Detailed Description
Hereinafter, the black raspberry extract of the present invention will be described. The black raspberry extract of the invention has the function of preventing cataract or inhibiting the development of cataract.
The black raspberry as a raw material of the black raspberry extract belongs to the genus Rubus of the family Rosaceae. In addition, the black raspberry extract of the present invention preferably uses an extract mainly from the fruit of black raspberry.
The method for extracting the black raspberry extract from the black raspberry fruit is not particularly limited, and solvent extraction, supercritical extraction, or the like may be used.
The extraction solvent used in the solvent extraction is not particularly limited, and water, an organic solvent, or a mixed solvent thereof may be used. As organic solvents, it is possible to use: alcohols such as methanol, ethanol, n-propanol, isopropanol, and n-butanol; glycols such as ethylene glycol and propylene glycol; ethyl acetate; acetone, and the like. These organic solvents may be used alone or in combination of two or more. In addition, an aqueous solvent such as water and an organic solvent may be used in combination.
Among them, ethanol and an ethanol aqueous solution obtained by mixing ethanol and water are preferable, and the concentration of ethanol in the ethanol aqueous solution is preferably 1 to 80 v/v% (EtOH), more preferably 10 to 60 v/v% (EtOH), and still more preferably 20 to 40 v/v% (EtOH).
When supercritical extraction is performed, carbon dioxide in a supercritical state is preferably used.
As an extraction method for performing solvent extraction, stirring extraction, immersion extraction, shaking extraction, reflux extraction, ultrasonic extraction, or the like can be used. Among them, stirring extraction is particularly preferable. The method of extraction by stirring is not particularly limited, and stirring with a stirrer, stirring with a masher, or the like can be performed.
In addition, in the case of extraction by stirring, extraction is preferably performed for several minutes to 60 minutes, more preferably for 30 minutes to 60 minutes, while stirring at a stirring speed of preferably 500 to 20000rpm, more preferably 1000 to 10000rpm, and even more preferably 6000 to 10000 rpm. After stirring, the mixture is preferably left to stand and extracted. More specifically, it is preferable to perform homogenization using a homogenizer at 6000 to 10000rpm for 30 to 60 minutes and then to perform extraction by standing for 24 hours. By allowing to stand and extracting, solid-liquid separation can be performed.
The temperature for solvent extraction may be room temperature or heated. The pressure at the time of solvent extraction may be normal pressure or increased pressure.
The extract obtained by solvent extraction may be subjected to a treatment such as filtration, if necessary.
Alternatively, the extract obtained by solvent extraction may be dried to obtain a powder. As the drying method, a known method can be used, and methods such as spray drying, vacuum drying, and freeze drying can be used.
As the black raspberry extract, a commercially available product or an existing product prepared as a liquid or powder can be used.
The black raspberry extract of the present invention can be incorporated into foods, beverages, and the like. Examples of the food include breads, noodles, snacks, processed meat products, processed fish products, frozen foods, jellies, ice creams, dairy products, and various seasonings. In addition to general foods, the composition can be incorporated into specific health foods, quasi drugs, health foods, and supplements. Examples of the beverage include soft drink water, milk beverage, alcoholic beverage, tea, black tea beverage, coffee, fruit juice beverage, carbonated beverage, mineral water, fruit and vegetable beverage, and the like.
In addition, the food and drink containing the black raspberry extract of the present invention may be prepared in the same form as oral preparations such as tablets, capsules, and syrups.
In the production of foods and beverages containing the black raspberry extract of the present invention, additives such as sweeteners, colorants, preservatives, thickeners, stabilizers, gelling agents, antioxidants, color developers, bleaching agents, emulsifiers, swelling agents, acidulants, glossing agents, and flavors, solvents, and oils may be added as needed within the range that does not impair the effects of the present invention. These additives may be used singly or in combination of two or more.
The ratio of the black raspberry extract to be incorporated into the food or drink may be appropriately adjusted depending on the purpose of use, and the ratio of the black raspberry extract to be incorporated into the food or drink is preferably 0.0001 to 80 wt%, more preferably 0.003 to 50 wt%, and still more preferably 0.005 to 30 wt%.
The black raspberry extract of the present invention is useful for the prevention or the progress inhibition of cataract.
Examples
The present invention will be described below with reference to examples, but the present invention is not limited thereto. Unless otherwise specified, the parts and% in the present example are based on weight. In examples and comparative examples, evaluation of antioxidant effect based on ORAC values and evaluation of expression induction of α -crystallin in lens cells were performed.
About a sample
(preparation of sample) as samples, extracts of three kinds of berries (black raspberry (rubus of rosaceae) (hereinafter, sometimes referred to as "BR"), blueberry (vaccinium of ericaceae) (hereinafter, sometimes referred to as "BB") and raspberry (rubus of rosaceae) (hereinafter, sometimes referred to as "RB")) and Blabina (registered trademark) as a powder preparation containing the extract of black raspberry were used.
(preparation of extract)
The extracts of black raspberry, blueberry and raspberry were obtained by the following method.
First, the fruits of black raspberry, blueberry and raspberry were preserved in a frozen state, respectively. Next, each fruit was thawed and cut. Black raspberry, blueberry and raspberry were weighed separately (about 100g), 5 times of 30 v/v% (EtOH) ethanol aqueous solution (about 500mL) was added, and extraction was performed with stirring using a blender at 8000rpm for 30 minutes. Further, the mixture was allowed to stand at room temperature for 24 hours, and then subjected to standing extraction, and filtered through a 150-mesh filter to obtain ethanol extracts of the black raspberry, the blueberry, and the raspberry, respectively. The following ORAC was measured in an ethanol-dissolved state, and therefore, the measurement was performed using an ethanol extract.
Incidentally, Blabina was dissolved in 30 v/v% (EtOH) so that Blabina was 9 mg/mL.
(preparation of extract)
Evaporating ethanol from the ethanol extracts of black raspberry, blueberry, raspberry and the ethanol solution of Blabina obtained by the above preparation of the extractive solutions respectively by using an evaporator to obtain extracts. It should be noted that the ethanol extracts of black raspberry, blueberry and raspberry contained 9mg/mL extract (extract).
Measurement and evaluation
(evaluation of antioxidation Effect based on ORAC value)
The ORAC values of the three resultant berry samples (black raspberry, blueberry, and raspberry ethanol extracts) were determined. The results are shown in table 1. Note that no ORAC value was measured for Blabina.
[ Table 1]
| Kind of sample | ORAC value (mu MTE/g) |
| BR | 62 |
| BB | 66 |
| RB | 49 |
The ORAC values of the three berry samples all showed high values as compared with those of the general antioxidant food (for the ORAC values of the general antioxidant food, see the above-mentioned non-patent document 4). Among the three berry samples, blueberry showed the highest value, but this was the result measured using the ethanol extract, and therefore, water-soluble components could not be involved.
In addition, since 0.1% or more of ethanol causes damage to cells, it is not appropriate to use an ethanol extract solution using a 30 vol% ethanol aqueous solution when measurement and evaluation are performed using cells. Therefore, in the following evaluation of the induction of α -crystallin expression in lens cells, samples each obtained by evaporating ethanol components from ethanol extract by an evaporator were used (see "preparation of extract" above).
(Induction of expression of alpha-crystallin in lens cells) lens cells (B-3, eye lens I (eye lens), human; ATCC No.; CRL-11421) were used as cells for experiments, and D-MEM medium supplemented with 10% FCS was used at 37 ℃ and 5% CO2The culture was carried out in an incubator under the conditions of (1). Immediately before the experiment, the cells were collected with a 0.25% Trypsin (Trypsin)/EDTA solution, and after trypan blue staining, the number of cells was calculated using an automatic cell counter (Invitrogen). Then, the cells were inoculated into 24-well plates or 96-well plates, and pre-cultured for 24 hours for the experiment.
The three berry extracts (black raspberry, blueberry, and raspberry) obtained in the preparation of the above extracts, and the ethanol-removed extract of Blabina were dissolved in ultrapure water to prepare about 11.5mg/mL, respectively, to obtain a sample. Subsequently, each sample was serially diluted (from 100000-fold to 10-fold), and the expression of α -crystallin-mRNA was analyzed by quantitative PCR.
The specific procedure of the quantitative PCR method is as follows. All RNA was extracted from the lens cells to which the sample was added using Trizol reagent (ambion). The extracted RNA was reverse-transcribed into cDNA according to the method of PrimeScript RT Master Mix (Takara), and amplified using SYBR Premix EX Taq II (Takara). In addition, 50. mu.L (25. mu.L of SYBR Green Mix (2X), 1. mu.L of cDNA, 2. mu.L of primer pair Mix (5 pmol/. mu.L of each primer)), and 22. mu. L H of primer pair Mix (each primer)) were used as PCR reaction solutions2O), the reaction is carried out under the following conditions: the reaction was carried out for 1 cycle at 95 ℃ for 30 seconds, and further for 50 cycles at 95 ℃ for 5 seconds and 60 ℃ for 30 seconds. Relative quantification was performed by the comparative Ct method (Δ Δ Ct method). Primers for various targets and housekeeping Genes (GAPDH) used in RT-PCR are shown in Table 2In (1).
[ Table 2]
After the serially diluted samples were added to lens cells, all RNAs were recovered at 1, 4, 8, and 12 hours, respectively. Compared with BB and RB, Blabina and BR both showed the induction of α -crystallin expression starting from 1 hour after addition even when diluted 100000-fold, and maintained high until 12 hours later (see fig. 1).
In order to maintain the lens for a long period of time, there are various factors, although the mechanism thereof has not been elucidated, but crystallins are constituent components of the lens, and the effect of α -crystallins as a molecular chaperone for preventing association of other β -and γ -crystallins is remarkable (see, for example, Andley up. crystallins in the Eye: Function and pathology. prog recording Eye res.2007 jan; 26 (1): 78-98.). Blabina, BR showed a higher tendency to induce the expression of alpha-crystallin than other berry extracts. In addition, in order to prevent cataract or inhibit the progression thereof, it is necessary to maintain the lens for a long period of time, and the above results suggest an effect of preventing cataract or inhibiting the progression thereof.
Claims (2)
1. A method for producing an alpha-crystallin expression inducer, which is an alpha-crystallin expression inducer comprising a black raspberry extract as an active ingredient, comprising:
freezing the fruit of the black raspberry;
a thawing step of thawing the fruit of the black raspberry;
a stirring step of adding 20 to 40 v/v% ethanol aqueous solution to the fruit of the black raspberry thawed in the thawing step and stirring the mixture; and
and an extraction step of standing and extracting the fruit of the black raspberry stirred by the stirring step and the ethanol aqueous solution.
2. Use of an expression inducer for alpha-crystallin comprising a black raspberry extract as an active ingredient, obtained by the production method according to claim 1, for the production of a medicament for preventing cataract or inhibiting the progression thereof.
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| JP2017154174A JP6535710B2 (en) | 2017-08-09 | 2017-08-09 | Method of producing expression inducer of α-Crystallin, cataract prevention / progression inhibitor containing expression inducer of α-Crystallin, and expression inducer of α-Crystallin |
| JP2017-154174 | 2017-08-09 | ||
| PCT/JP2018/006354 WO2019030954A1 (en) | 2017-08-09 | 2018-02-22 | Black raspberry extract, cataract prevention/progression inhibitor including black raspberry extract, food and beverage, production method for black raspberry extract, cataract prevention method or progression inhibiting method using black raspberry extract |
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| EP1683805A1 (en) * | 2003-10-24 | 2006-07-26 | Meiji Seika Kaisha Ltd. | Novel inhibitor for advanced glycation endproduct formation and aldose reductase inhibitor |
| CN102697893A (en) * | 2012-05-22 | 2012-10-03 | 辽宁大学 | Application of black raspberry extract (BRBE) in preparation of drugs for treating gastric cancer |
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| US5128365A (en) * | 1989-07-19 | 1992-07-07 | The Trustees Of Columbia University In The City Of New York | Compounds having glutathione peroxidase activity and use thereof |
| JPH06199692A (en) * | 1992-03-06 | 1994-07-19 | Yunie:Kk | Agent for amelioration and treatment of cataract |
| US7208181B1 (en) * | 2002-06-12 | 2007-04-24 | The United States Of America, As Represented By The Secretary Of Agriculture | Isolation of polyphenolic compounds from fruits or vegetables utilizing sub-critical water extraction |
| CN1884467A (en) * | 2006-05-25 | 2006-12-27 | 任伟庆 | Black raspberry juice wine and process for preparation |
| KR100877371B1 (en) * | 2007-03-30 | 2009-01-09 | 주식회사 로제트 | Compositions for Protecting Eyesight |
| JP2013018734A (en) * | 2011-07-09 | 2013-01-31 | Nof Corp | Hyaluronic acid production promoter |
| CN102613631B (en) * | 2012-03-31 | 2014-02-12 | 贵州北极熊实业有限公司 | Black raspberry beverage and making method thereof |
| KR101978147B1 (en) * | 2012-11-15 | 2019-05-15 | (주)아모레퍼시픽 | Perfume composition for expressing the fragrance of Rubus coreanus Miquel |
| KR20160116487A (en) * | 2015-03-30 | 2016-10-10 | 전라북도 고창군 | Functional Gel Using Unripened Black Raspberry and Manufacturing Method Thereof |
| JP6643147B2 (en) * | 2016-03-04 | 2020-02-12 | 株式会社Ｓｕｂａｒｕ | Casting apparatus and casting method |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1683805A1 (en) * | 2003-10-24 | 2006-07-26 | Meiji Seika Kaisha Ltd. | Novel inhibitor for advanced glycation endproduct formation and aldose reductase inhibitor |
| CN102697893A (en) * | 2012-05-22 | 2012-10-03 | 辽宁大学 | Application of black raspberry extract (BRBE) in preparation of drugs for treating gastric cancer |
Non-Patent Citations (4)
| Title |
|---|
| Anti-cataractogenesis Mechanisms of Curcumin and a Comparison of its Degradation Products: an In Vitro Study;Jiahn-Haur Liao et al.;《AGRICULTURAL AND FOOD CHEMISTRY》;20160223;第1-28页 * |
| Blackberry, raspberry and black raspberry polyphenol extracts attenuate angiotensin II-induced senescence in vascular smooth muscle cells;Rafaela G. Feresin et al.;《Food & Function》;20160726;第1-13页 * |
| Green tea flavanols protect human γB-crystallin from oxidative photodamage Susmitnarayan Chaudhury, Pritam Roy, Swagata Dasgupta;Susmitnarayan Chaudhury et al.;《BIOCHIMIE》;20170308;第1-36页 * |
| Inhibitory effect of anthocyanins and colored rice on diabetic cataract formation in the rat lenses;Yasujiro Morimitsu et al.;《International Congress Series》;20021231;第503-508页 * |
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