JPS63179824A - Antimutagnetic agent - Google Patents

Abstract

PURPOSE:To obtain an antimutagenic agent for food stuff, containing kaempferol as an active ingredient, having high safety and showing excellent effect. CONSTITUTION:The mutagenicity of a mutagen is inhibited by 50%, by adding kaempferol which is a sort of flavonoid to three times amount of the mutagen. The kaempferol shows strong inhibitory effect to a mutagen, especially heterocyclic amines existing in scorched foods. The kaempferol exists in higher plants in nature and may be used by extracting and isolating from a synthetic substance or plant or used as a plant extract. The plant includes Carthamus tinctorius, Japanese pagoda tree, bitter orange, tangerine, Japanese green gentian, mulberry tree, licorice, etc.

Description

[Detailed Description of the Invention] [Industrial Application Field] The present invention provides a highly safe and highly effective product containing kaempferol, which is a type of flavonoid.
Concerning antimutagenic agents for food.

[Prior Art] Currently, cancer (malignant neoplasm) is the leading cause of death in Japan, and it is believed that most human cancers are caused by carcinogenic factors in the environment. It is being Among the carcinogenic factors that exist in this environment, the foods and drinks we consume on a daily basis are considered the most important, and there is an overwhelming amount of epidemiological data that supports this view.

In recent years, the Neems test, which uses histidine requirement as an index, has been established as a means of predicting the carcinogenicity of environmental factors such as foods and drinks and medicines (Ames, B.

N. et al.: Mutation Res.
Vol 31.347. '75).

Many research institutions have found a high correlation of 80-90% between the mutagenicity and carcinogenicity of compounds tested using this method (McCann).

J, et al.: Proc, Natl, Aea
d, Sci, (U, S, A,).

Vol, 72.5135. ’75).

Therefore, eliminating mutagenicity is expected to have a considerable effect on protecting humans from the risk of carcinogenesis. Also,
Moreover, mutations occur as a result of damage to DNA, which is genes, and even if they do not lead to cancer, the effects of mutagens on human health cannot be underestimated. This is why food and drink products that are not mutagenic are desired. To date, the mutagens present in food and beverages include nitrosamines produced by the reaction of nitrous acid and secondary amines, heterocyclic amines present in burnt food, and mycotoxins such as aflatoxin. Many of them have been proven to be carcinogenic. However, food and drinks are things we ingest on a daily basis, and we often ingest these mutagens without realizing it. In particular, many of the heterocyclic amines present in burnt food are highly mutagenic (see Table 1).
nmental Muta3ens and Carc
ino-gens), University of Tokyo Press
(Excerpt from O Press, Tokyo), p. 7, 1981).

(Left below) Table 1 Mutagenicity of heterocyclic amines present in burnt food *Abbreviation Isolated source ): (*Mutation activity M
eIQ Grilled Sardine 6610
00IQ Grilled Sardine 43
3000MeIQx Yakiniku
1.45000 Trp-P-2) Heating of liptophan 104000 Product (hereinafter the same) Orn-P-1 Ornithine 56800
Glu-P-1 glutamic acid 4.900
0Trp-P-1 tryptophan 390
00Glu-P-2 glutamic acid 19
00AαC soybean globulin 300MeA
αC soybean globulin 200Lys-P-
1 Lysine 86Phe-P-1
Phenylalanine 41>:ζMeIQ
: 2-amino-8,4,-dimethylimidazo[4-,
5-f]quinoline, IQ: 2-amino-3-methylimidazo[4,5-f]quinoline, MeIQx: 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline, Trp-P- 2: 3-amino-1-methyl-
5H-pyrido[4,3-b]indole, 0rn-P-
1: 4-amino-6-methyl-1,H-2,5,10
, 10b-tetraazafluoranthene, Glu-P-1
,: 2-amino-6-methyldipyrido [1,2-a:
3°,2“-d]imidazole, Trp-P-1: 3
-Amino-1゜4-dimethyl-5H-pyrido[4,3-
b] Indole, Glu-P-2: 2-aminodipyrido[1°2-a:3',2'-d]imidazole, Aa
C: 2-amino-9H-pyrido[2,3-b]indole, MeAαC: 2-amino-8-methyl-9H-pyrido[2,3-b]indole, Lys-P-1: 3,4
- cyclopentinoviride [3,2-a]carbazole,
Phe-P-1: 2-amino-5-phenylpyridine *>: ζ Salmonella typhimurium (Sa1mon
Results of an experiment using the TA98 strain (A typhimurium) in the presence of rat liver homogenate (S9 mix). Number of revertant colonies generated per 1 μg of sample.

(The following is a blank space) The present invention aims to improve the safety of food and drink products by eliminating or attenuating their mutagenicity. Research in this field has produced many results to date, and flavonoids are also present in benzo [a], which is present in cigarette smoke and automobile exhaust gas.
] It was known that it exhibits an inhibitory effect on polycyclic aromatic hydrocarbon mutagens such as pyrene (Carcinogenesis, Vol. 4, pp. 1631-1637, 1983).

[Problems to be Solved by the Invention] In order to achieve the above object, the present inventors have made extensive research efforts and have found that kaempferol is a mutagen in foods, particularly in burnt food. We have discovered that this product has a strong inhibitory effect on existing heterocyclic amines.

The use of the above ingredients to produce non-mutagenic foods and drinks, or to eliminate the mutagenicity of foods and drinks, is completely new.

[Means for Solving the Problems] That is, the present invention is an antimutagenic agent for food, which is characterized by containing kaempferol as an active ingredient.

The configuration of the present invention will be explained in detail below.

Kaempferol used in the present invention naturally exists in higher plants. In the present invention, kaempferol may be synthesized or extracted and isolated from plants, etc., or may be used as a plant extract. Examples of such plants include safflower, apricot, Japanese daisy, tangerine, Japanese tangerine, mulberry, daylily, scutellaria, mugwort, kudzu, and the like.

According to Kuwai, MelQx detected in beef and chicken grilled on a gas stove and beef grilled on an electric hot plate was 0.5 ng per gram of grilled meat.
, 2.1 ng and 0.3 ng (Environmental Mutagen Research, Vol. 7, No. 1, pp. 35-41, 1985
).

Although MeIQx alone cannot explain all of the mutagenicity of these grilled meats, adding kaempferol at three times the amount of these mutagens suppressed the mutagenicity by 50%. Can be done. Assuming that 10 times more mutagens were present in the grilled meat, and the amount of yakiniku eaten at one time was 1008, the amount of mutagen ingested would be 2.1 μg for chicken grilled on a gas stove. , the amount of kaempferol required for this is 6.3
It becomes μg. If 100% suppression is desired, it is sufficient to use 10 times this amount, that is, 63Pg.

This is only 0.63 ppm in weight ratio to meat.
It can be said that a sufficient effect can be obtained with an extremely small amount. [Effects of the Invention] Next, the present invention will be explained in detail with reference to test examples.

(Test example) ■Measurement method of mutagen suppression effect (1) Method: Pre-incubation method (cedar wood, long legs:
Chemical Mutadiens (ChemiCalMut)
Gens), Vol. 6, p. 41, 1981).

(11) Bacterial strain used: histidine auxotrophic Salmonella typhimurium (Salmonella typhimurium)
rium ) TM01 strain (hereinafter abbreviated as TM98).

(il+) Sample preparation 1) For kaempferol and other flavonoids (standard): These are dimethyl sulfoxide (hereinafter referred to as
Dissolve in DMSO (abbreviated as DMSO).

2) In the case of safflower extract: Safflower was extracted with ethyl acetate, and the extract was evaporated to dryness to obtain an ethyl acetate extract of safflower. Furthermore, the extract obtained by extracting safflower with hot water was further extracted with acetone, and the obtained extract was purified by activated carbon treatment to obtain a safflower extract. These were also dissolved in DMSO.

(iv) Measurement of mutagenicity Each sample 5 obtained in 1) and 2) of (iil) above
Add nicotinamide adenine dinucleotide (NADH) to 0 μl and 500 μm of 89 mix (9000×g supernatant of rat liver homogenate in which the drug-metabolizing enzyme system was induced with phenoparbital and 5,6-benzoflavone).
, nicotinamide adenine dinucleotide phosphate (NA
DPH) and coenzymes such as glucose-6-phosphate (06P), magnesium ions (Mg2+), etc.), 100 μl of TA98 strain preculture solution and 50 μm
mutagens (Trp-P-1, Glu-P-1, IQ
etc.). After shaking this mixture at 37°C for 20 minutes, it is mixed with 2 ml of dissolved soft agar containing trace amounts of histidine and biotin, and spread on a minimum glucose agar medium. After standing at 37°C for 48 hours, count the number of revertant colonies that do not require histidine on the medium. In addition, the mutagenicity suppressing effect is calculated based on the number of colonies generated on the medium containing only the mutagen as 100, and the extent to which the number of colonies decreased when kaempferol was added is expressed as a percentage of the above control. . The calculation formula is shown below.

A=l(B-C)÷(D-C)l Number of Colonies II, Results (i) Mutagen Suppression Effect of Kaempferol The results are shown in the table below.

Flavonoid 50% inhibition amount (ng) Quercetin 390 450 150 Kaempferol 130 140 100 Phisetin 5
70 1350 1320 Morin 1.1
50 2390 2120 Myricetin 35
0 350 320*: Addition amount is Trp-P-1:
42 ng, Glu-P-C 40 ng.

IQ: 10 ng.

The values in the table indicate the amount of flavonoid required to suppress the mutagenicity of the mutagen by 50%. Kaempferol has a stronger inhibitory effect than other flavonoids, and when targeting Trp-P-1 and Glu-P-1, it has been shown to be effective at about one-third the amount of quercetin, for example. Ta.

(it) The results of the mutagenic inhibitory effect of safflower extract are shown in Figure 1.
and 2. Here, as a mutagen, Tr
p-P-1 (42 ng) was used. The 50% inhibition amount was <Img for Safflower ethyl acetate extract and 2.7 mg for Safflower extract.

[Example] EXAMPLES The present invention will be explained in more detail with reference to Examples below.

The present invention is not limited to these.

Example 1 Orange juice concentrate 190g Sugar 260g Citric acid 2g Flavoring
4.5g Kaempferol 3mg Dilute the total amount with 3mg pure water
Adjust to 000g.

Orange juice was obtained with the above recipe.

Example 2: Mixed fruit drink Passion juice 200g Orange Piure - 1008 Apple juice concentrate 10g Sugar 100g Citric acid 0.4g Safflower extract 0.688 The total volume was made up to 1000ml with pure water.

A mixed fruit drink was obtained with the above formulation.

[Brief explanation of the drawing]

Figures 1 and 2 show safflower ethyl acetate extract,
This shows the inhibitory effect of safflower extract on Trp-P-1. The horizontal axis is the amount of extract added per plate (mg: milligrams), and the vertical axis is the number of reverted mutation colonies that occurred without adding the extract. It is shown as a ratio to the number of colonies on the control plate (this is taken as 100).

Claims (1)

[Claims] An antimutagenic agent for food, characterized by containing kaempferol as an active ingredient.