JP2007137794A - Wasabi leaf component composition and food and drug containing the same - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a Wasabi (Japanese horseradish) leaf component composition which exhibits strong detoxication metabolizable enzyme induction activity to thereby improve the detoxication metabolizable function, further inhibits nephropathy caused by drugs and diabetes, and resultingly reduces the risk of cancer and life style related diseases, and food and drink and a drug containing the same. <P>SOLUTION: The Wasabi leaf composition comprises a Wasabi leaf component composed of Wasabi leaves, horseradish leaves or their extract, and exhibits the action of inducing activation of the phase II detoxication metabolizable enzyme, and exhibits the action of inhibiting nephropathy. The food and drink and the drug contain this composition as the major component. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to a wasabi leaf component composition that induces the activity of a second-phase detoxifying metabolic enzyme in a living body, and further exhibits a renal disorder inhibitory action, a food and drink containing the composition, and a pharmaceutical product. Wasabi, which activates phase II detoxification metabolic enzymes in metabolism, further increases the amount of these enzymes, has an effect of inhibiting the degradation of the enzyme itself, or reduces renal oxidative stress by reducing in vivo oxidative stress. The present invention relates to a leaf component composition, a food or drink containing the composition, and a pharmaceutical product.

  Today's living environment is flooded with harmful substances that have harmful effects on living organisms. Typical examples are dioxins, mercury, lead, cadmium, and various pesticides. These harmful substances pollute the soil, air, and seawater, and enter the living body through food and drinking water.

  In Japan, which is surrounded by the sea on all sides, it is estimated that 40% of animal protein is taken from seafood. Hazardous substances discharged into the environment are likely to accumulate in aquatic products at high concentrations through the food chain, so it can be said that there is a high risk of harmful substances accumulating in the body (intermediate study on the effects of endocrine disruptors on agricultural, forestry and fishery products) Report April 1999, Ministry of Agriculture, Forestry and Fisheries).

  Moreover, cadmium contamination of food in Japan is serious. According to the results of the 2001 survey by the Ministry of Health, Labor and Welfare, the intake of cadmium from Japanese daily food is 4.102 μg / kg / week, and FAO / WHO is It reaches about 60% of the provisional tolerable intake set (Q & A on “Cadmium in foods” 2004 Ministry of Health, Labor and Welfare).

  Furthermore, dioxins taken into the body, mercury, lead, cadmium, various pesticides and other harmful substances, and the accumulation of these metabolites directly cause drug-induced kidney damage, and diabetic nephropathy It is an indirect cause.

  According to the 2002 survey of diabetes by the Ministry of Health, Labor and Welfare, it is estimated that there are approximately 7.4 million people who are strongly suspected of having diabetes, and 16.2 million people who cannot deny the possibility of diabetes. In addition, diabetic nephropathy causes more than 11,000 artificial dialysis patients every year (2002 Diabetes Survey (Bulletin 2002), Ministry of Health, Labor and Welfare).

The hyperglycemic state in diabetes causes non-enzymatic glycation of proteins in the living body and produces advanced glycation end products (hereinafter referred to as AGE) through a complex reaction.
AGE damages the glomerular endothelial cells and mesangial cells of the kidney, causing diabetic nephropathy.

  In general, the living body is equipped with a two-phase detoxification mechanism for metabolizing and detoxifying harmful substances. In the first phase, Ah receptors to which harmful substances are bound bind to the transcription region of the gene and increase the transcription amount of detoxifying metabolic enzymes such as CYP1A1 and CYP1A2. These first phase detoxifying metabolic enzymes oxidatively modify harmful substances and form reaction intermediates. These reaction intermediates may form adducts with DNA and proteins, and may exhibit acute toxicity and carcinogenicity.

  In the second phase, the transcription factor Nrf2 is released from the sensor molecule Keapl by exposure to the above reaction intermediate and harmful substances, and the second phase detoxification metabolic enzyme is induced. Phase II detoxifying metabolic enzymes carry out the conjugation reaction of hydrophilic functional groups of harmful substances and excrete them out of the body.

  If the activity of such detoxifying metabolic enzymes is further improved or increased, the second phase detoxifying metabolic reaction is activated and the risk of poisoning and carcinogenesis due to harmful substances is reduced.

  In addition, activation of heme oxygenase (hereinafter referred to as “HO”), which is an oxidative stress responsive enzyme, and glutathione S-transferase (hereinafter referred to as “GST”) conjugated with a peroxidation product cause the accumulation of peroxidation products. It is possible to prevent and improve lifestyle-related diseases such as arteriosclerosis and diabetes.

  Serum thymus factor (FTS nonapeptide) (International Publication Number: WO2002 / 017965) and a compound that induces GST activity (International Publication Number: WO95 / 09866) are known as compounds that induce the activity of detoxifying metabolic enzymes in the body. However, it is difficult to take these substances on a daily basis.

  6-MSITC (6-methylsulfinylhexyl isothiocyanate) (Japanese Patent Application No. 11-240823), which is specifically contained in the roots of this horseradish, is a natural product that promotes the activation of detoxifying metabolic enzymes such as GST and prevents carcinogenesis. ), Sulforaphane (4-methylsulfinylbutyl isothiocyanate) (Japanese Patent Laid-Open No. 2005-60229) contained in broccoli, and broccoli sprout and the like are marketed as foods with high antioxidant activity.

  Examples of drugs that suppress drug-induced renal injury include vascular endothelial growth factor / vascular permeability factor (VEGF / VPF) (Japanese Patent Laid-Open No. 2000-169390), xanthine derivatives (International Publication Number: WO 98/57644), MK family Proteins (International Publication No .: WO98 / 40095) and the like belonging to are disclosed.

  As a drug for suppressing diabetic nephropathy, a TGF-β inhibitor that suppresses fibrosis of kidney tissue, a CTGF inhibitor (JP 2004-131444), and a pyridinium compound that decomposes AGE (JP 2002-275158) In addition, gluconic acid or a salt thereof (Japanese Patent Laid-Open No. 9-194377) that suppresses calcification of kidney tissue has been devised.

  Although the aforementioned sulforaphane exhibits a strong GST-inducing activity, its action is still insufficient due to its low content and stability. Moreover, since many of the renal disorder inhibitors currently devised are pharmaceuticals or synthetics, it is difficult to take them on a daily basis.

Nowadays, toxic substances are inundated and forced to be exposed to oxidative stress environment. Stronger and more easily developed detoxification metabolic enzyme activity induction / kidney disorder suppression composition, or food / beverage products and medicines containing these are strongly developed. It is desired.
International Publication Number WO2002 / 017965 International Publication Number WO95 / 09866 Japanese Patent Application No. 11-240823 JP2005-60229 JP 2000-169390 International Publication Number WO98 / 57644 International Publication Number WO98 / 40095 JP2004-131444 JP 2002-275158 A JP-A-9-194377 Interim report on the environmental problem study meeting of endocrine disrupting substances to agricultural, forestry and fishery products 1999 Ministry of Agriculture, Forestry and Fisheries Q & A on “Cadmium in Foods” 2004 Ministry of Health, Labor and Welfare 2002 Survey on Diabetes Mellitus (Preliminary Report) 2002 Ministry of Health, Labor and Welfare

  Therefore, the object of the present invention is to exhibit a strong detoxification metabolic enzyme-inducing activity, thereby improving the detoxification metabolic function, further suppressing renal damage caused by drugs and diabetes, and consequently reducing the risk of cancer and lifestyle-related diseases. Then, it is providing the wasabi leaf component composition which improved the fault which exists in the above-mentioned well-known technique, the food-drinks containing this, and a pharmaceutical.

  In order to solve the above-mentioned problems, the wasabi leaf component composition of the present invention includes a wasabi leaf component comprising the present wasabi leaf, horseradish leaf, or an extract thereof, and is a second-phase detoxifying metabolic enzyme in vivo. It exhibits an activity-inducing action.

  Furthermore, in order to solve the above-described problems, the wasabi leaf component composition of the present invention includes the wasabi leaf component, the wasabi leaf component composed of the wasabi leaf, the horseradish leaf, or an extract thereof, and reduces oxidative stress in the living body. It is characterized by exhibiting an action to suppress kidney damage.

  Furthermore, in order to solve the above-mentioned problem, according to the food and drink of the present invention, the wasabi leaf according to claim 1 or 6 containing the wasabi leaf component comprising the present wasabi leaf, horseradish leaf, or an extract thereof. It contains an ingredient composition as a main component.

  Furthermore, in order to solve the above-mentioned problem, according to the pharmaceutical product of the present invention, the wasabi leaf component composition according to claim 1 or 6, comprising the wasabi leaf component comprising the present wasabi leaf, horseradish leaf, or an extract thereof. It is characterized by containing a product as a main component.

  The present invention contains wasabi leaves, horseradish leaves, or wasabi leaf components composed of these extracts as active ingredients, and ingests these as foods and drinks or as pharmaceuticals, so that the detoxifying metabolic enzyme activity in vivo can be obtained. It has the effects of inducing and inhibiting kidney damage and reducing the risk of cancer and lifestyle-related diseases.

  Hereinafter, the present invention will be described in detail.

  The raw material used in the present invention is a wasabi leaf component, a horseradish leaf, a horseradish leaf component made of these extracts. That is, plants as raw materials used in the present invention are wasabi and horseradish, and the part to be used is leaves. These two kinds of plants are used alone or in a mixture at an arbitrary ratio.

  The above-mentioned wasabi leaf component is obtained by pulverizing or grated this wasabi leaf or horseradish leaf, extraction treatment with a solvent, drying treatment, or a combination treatment thereof. That is, the active ingredients of the above-mentioned wasabi leaves and horseradish leaves are obtained by physical means such as pulverization or cutting of plants, grated, or water, methanol, ethanol, acetone, ethyl acetate, diethyl ether, dichloromethane, dichloroethane, etc. It can be obtained by a solvent extraction means, a drying means such as hot air drying, freeze drying, spray drying, or a combination of these treatments.

  The wasabi leaf component according to the present invention thus obtained contains polyphenols, tannins, indoles, organic acids, chlorophyll and the like as active ingredients. In addition, as the above polyphenols, flavonoids such as apigenin, luteolin, kaempferol and their glycosides, multimers, phenylpropanoids such as ferulic acid and sinapinic acid, their glycosides, multimers, these A mixture is mentioned.

  The wasabi leaf component of the present invention does not include isothiocyanates. Therefore, isothiocyanates are not involved in the detoxification metabolic enzyme activity induction and the renal disorder inhibitory action in the present invention.

  The above-mentioned wasabi leaf component composition according to the present invention can be ingested alone, but can also be used by adding to various foods, beverages, medicines and quasi drugs. Specifically, liquid foods such as soft drinks, tea beverages, drinks, alcoholic beverages, solid foods such as confectionery, cooked rice, breads, noodles, seasonings, powders, granules, capsules, tablets, etc. It can be added to other pharmaceuticals and quasi drugs.

  Such a wasabi leaf component composition according to the present invention is in vivo as it is, or added to foods and drinks, and further added to drugs and quasi-drugs for ingestion. In two-phase foreign body metabolism, GST, quinone reductase (hereinafter referred to as “QR”), N-acetyltransferase (hereinafter referred to as “NAT”), HO and other second phase foreign body metabolic enzymes are activated, and further, these enzymes are increased in action. In addition, it exhibits an inhibitory effect on the degradation of the enzyme itself, and further exhibits an inhibitory effect on renal damage by reducing oxidative stress in vivo.

Extraction of this wasabi leaf This wasabi leaf extract was prepared. 16 kg of 50% ethanol was added to 4 kg of this wasabi leaf finely cut and extracted for 1 hour with stirring. The extract was filtered, concentrated with an evaporator, and freeze-dried, and then pulverized with a food mill to obtain 70 g of a brown powder. This was subjected to the test as this wasabi leaf extract.

  The results of subjecting this wasabi leaf extract to high performance liquid chromatography (hereinafter "HPLC") are shown in FIG. Several kinds of flavonoids and phenylpropanoids were detected from this wasabi leaf.

Extraction of horseradish leaves A horseradish leaf extract was prepared. 16 kg of hot water was added to 4 kg of finely cut horseradish leaves, and extraction was performed for 15 minutes while stirring and heating. The extract was filtered, concentrated with an evaporator, and then freeze-dried, and pulverized with a food mill to obtain 85 g of a brown powder. This was subjected to testing as a horseradish leaf extract.

  The results of subjecting the horseradish leaf extract to HPLC are shown in FIG. Several kinds of flavonoids were detected from horseradish leaves.

Analysis of detoxifying metabolic enzyme-inducing activity of wasabi leaf extract The wasabi leaf extract was obtained by the method described in Example 1, and the horseradish leaf extract was obtained by the method described in Example 2.

  36 ICR mice (male, 6 weeks old) were preliminarily raised in a stainless steel individual cage for 1 week, then water (12 animals), 3% horseradish leaf extract (12 animals), 3% horseradish leaf extract (12 animals) Were allowed to ingest each group for 1 week. The temperature of the animal room was maintained at 24 ± 1 ° C., and the animals were reared at a humidity of 50%, a light period of 08: 00-20: 00, and a dark period of 20: 00-08: 00. Blood was killed by exsanguination under ether anesthesia, and the liver tissue was removed, rapidly frozen in liquid nitrogen and stored at -80 ° C.

  The obtained liver was perfused with 1.15% KCl and stored at −80 ° C. until use. To this liver, 4 volumes of KCl was added and homogenized with a homogenizer, 1.5 ml was taken and centrifuged at 750 × g for 10 minutes. Subsequently, the supernatant was centrifuged at 10,000 × g for 10 minutes, the supernatant was further centrifuged at 100,000 × g for 70 minutes, and the obtained supernatant was subjected to enzyme activity measurement.

Protein measurement The amount of protein in the supernatant was measured using a BCA protein assay reagent kit (PIERCE), 200 μl of the mixed reagent for measurement was added to 10 μl of the supernatant, incubated for 30 minutes, and the absorbance at 562 nm was measured to measure the protein. This was done by quantifying the amount.

GST activity measurement In principle, the method of WHHabig et al. (J. Bio. Chem. 249, 713, 1974) was followed. That is, 0.1 ml of the sample obtained above was added to a mixed solution of 0.5 ml of 0.2 M phosphate buffer (PH 6.5), 0.1 ml of 10 mM reduced glutathione aqueous solution and 0.2 ml of purified water, and stirred. Thereafter, 100 μl of 10 mM CDNB (1-chloro-2,4-dinitrobenzene) was added to start the reaction, and then the absorbance at 340 nm was measured every 15 seconds for 3 minutes, and the activity was measured from a calibration curve.

The GST activity value was determined by the following arithmetic formula.
GST activity value (U / mg protein) = (absorbance at 340 nm / 3 × 10 × 100) / (9.6 × protein amount)

Measurement of QR activity In principle, the method of C. Lind et al. (Method in enzymology 186, 1990) was followed. That is, 150 mM Tris-HCl containing 0.24% Triton X-100 and distilled water are mixed so as to have a ratio of 1: 1.6, and are heated in a constant temperature bath at 37 ° C. 2.6 ml of the above solution and 0.1 ml of 15 mM NADPH, 0.1 ml of 2.3 lmM cytochrome c, 0.1 ml of sample are placed in a glass cell. A glass cell is set on a spectrophotometer filter adjusted to 30 ° C. and allowed to stand for 1 minute. 0.1 ml of 0.3 mM menadione was placed in the cell and stirred, and the activity was measured by measuring the absorbance at 550 nm every 20 seconds for 3 minutes.

QR activity was determined by the following arithmetic formula.
QR activity (U / mg protein) = {ΔA (sample) −ΔA (blank)} × dilution rate / ε × 10 ⁻³ × protein amount (in sample added to reaction solution)
Absorbance change for 1 minute at ΔA = 550 nm
ε = 18500 (Molar extinction coefficient of reduced cytochrome c)

  The measurement results are shown in FIGS. FIG. 3 shows that the GST activity in mice was significantly increased by administration of the present wasabi leaf extract and the horseradish leaf extract according to the present invention.

  It can be seen from FIG. 4 that mouse QR activity was increased by administration of the wasabi leaf extract and the horseradish leaf extract according to the present invention.

  To mice bred in the same manner as in Example 3, iron nitrilotriacetic acid was intraperitoneally administered, and the amount of kidney TBARS and serum creatinine after 3 hours were measured.

  FIG. 5 and FIG. 6 show that administration of this wasabi leaf extract and horseradish leaf extract reduced the TBARS value and the amount of creatinine in mice and suppressed drug-induced renal injury.

Production of tablet with this wasabi leaf extract 3 parts of this wasabi leaf extract prepared in Example 1 was doubled with fructose and tableted to obtain a tablet having a diameter of 1.0 g and a diameter of 7 mm.

Preparation of horseradish leaf green juice The dried horseradish leaf was pulverized by a pulverizer and then passed through a 100 mesh sieve to form a fine powder. 50 g of this horseradish leaf fine powder and commercially available barley young leaf powder were weighed and mixed to obtain a raw powder of horseradish leaf green juice. 3 g of this was weighed and mixed with 100 ml of water for drinking.

  The present invention contains wasabi leaves, horseradish leaves, or wasabi leaf components composed of these extracts as active ingredients, and ingests these as foods and drinks or as pharmaceuticals, so that the detoxifying metabolic enzyme activity in vivo can be obtained. It has the effects of inducing and inhibiting kidney damage and reducing the risk of cancer and lifestyle-related diseases. Therefore, the present invention is highly applicable in the medical field.

It is the graph which showed the result of having used this wasabi leaf extract for HPLC. It is the graph which showed the result which used the horseradish leaf extract for HPLC. It is the graph which showed the GST activity induction effect of this wasabi leaf extract and a horseradish leaf extract. It is the graph which showed the QR activity induction effect of this wasabi leaf extract and a horseradish leaf extract. It is the graph which showed the TBARS reduction effect | action of this wasabi leaf extract and a horseradish leaf extract. It is the graph which showed the creatinine reduction effect of this wasabi leaf extract and a horseradish leaf extract.

Claims (17)

  A wasabi leaf component composition comprising a wasabi leaf component comprising this wasabi leaf, horseradish leaf, or an extract thereof, and exhibiting an activity inducing activity of a second phase detoxifying metabolic enzyme in vivo.   The wasabi leaf component composition according to claim 1, wherein the wasabi leaf component contains one or more selected from the group of polyphenols, tannins, indoles, organic acids, and chlorophyll as an active ingredient.   The wasabi leaf component composition according to claim 2, wherein the polyphenols are flavonoids and their glycosides and multimers, phenylpropanoids and their glycosides and multimers, or a mixture thereof. object.   The wasabi leaf component composition according to claim 1, wherein the wasabi leaf component is obtained by pulverizing or grated this wasabi leaf or horseradish leaf, extraction with a solvent, drying treatment, or a combination thereof. .   The detoxifying metabolic enzyme according to claim 1, wherein glutathione S-transferase, quinone reductase, N-acetyltransferase, heme oxygenase, UDP-glucuronosyltransferase, sulfotransferase, acetyltransferase, catechol 0-methyltransferase and acids: CoA ligase and The wasabi leaf component composition according to claim 1, which is one or more selected from the group of acyl-CoA: amino acid N-acyltransferases.   A wasabi leaf component composition comprising a wasabi leaf component comprising the present wasabi leaf, horseradish leaf, or an extract thereof, and exhibiting a renal injury inhibitory action by reducing oxidative stress in vivo.   The wasabi leaf component composition according to claim 6, wherein the wasabi leaf component contains one or more species selected from the group of polyphenols, tannins, indoles, organic acids, and chlorophyll as an active ingredient.   The wasabi leaf component composition according to claim 7, wherein the polyphenols are flavonoids and their glycosides and multimers, phenylpropanoids and their glycosides and multimers, or a mixture thereof. object.   7. The wasabi leaf component composition according to claim 6, wherein the wasabi leaf component is obtained by pulverizing or grated this wasabi leaf or horseradish leaf, extraction with a solvent, drying treatment, or a combination thereof. .   6. The renal injury according to claim 6, wherein the renal injury is a drug-induced renal injury caused by a heavy metal comprising cadmium, hexavalent chromium, mercury, lead, arsenic, selenium, fluorine, boron, cyanide, or a compound containing these. 6. Wasabi leaf component composition according to 6.   The wasabi leaf according to claim 6, wherein the renal disorder is a drug-induced renal disorder caused by dioxins composed of polychlorinated dibenzo-para-dioxin (PCDD), polychlorinated dibenzofuran (PCDF), or coplanar PCB. Ingredient composition.   7. The renal disorder according to claim 6, wherein the renal disorder is carbon tetrachloride, 1,2-dichloroethane, 1,1-dichloroethylene, cis 1,2-dichloroethylene, 1,3-dichloropropene, dichloromethane, tetrachloroethylene, 1,1,1-trichloroethane. The wasabi leaf component composition according to claim 6, which is a drug-induced nephropathy caused by volatile organic compounds consisting of 1,1,2-trichloroethane, trichloroethylene, benzene, or analogs of these compounds. In claim 6, the renal disorder is copper agent, sulfur agent, polyhaloalkylthio agent, organic chlorine agent, organic phosphorus agent, benzimidazole agent, dicarboximide agent, carboxyamide agent, acylalanine agent, EBI agent, The wasabi leaf component composition according to claim 6, which is a drug-induced nephropathy caused by an agrochemical comprising a strobilurin agent, a carbamate agent, a synthetic pyrethroid agent, a nereistoxin agent, a BT agent, or a neonicotinoid agent.   The wasabi leaf component composition according to claim 6, wherein the renal disorder is diabetic nephropathy.   The wasabi leaf according to claim 14, wherein the diabetic nephropathy is caused by increased polyol metabolic activity, final glycation product (AGE), glomerular hyperfiltration, or protein kinase C activation, respectively. Ingredient composition.   7. A food or drink comprising the wasabi leaf component composition of claim 1 or 6 containing the wasabi leaf component comprising the present wasabi leaf, horseradish leaf, or an extract thereof. A pharmaceutical product comprising the wasabi leaf component composition of claim 1 or claim 6 containing the wasabi leaf component comprising the present wasabi leaf, horseradish leaf, or an extract thereof.

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