JPS63211226A - Mutagenicity inhibitor - Google Patents

Abstract

PURPOSE:To obtain a mutagenicity inhibitor containing a specific organo- germanium compound as an active ingredient. CONSTITUTION:A mutagenicity inhibitor containing a compound expressed by the formula [R1-R3 are H, lower alkyl or (substituted)phenyl] as an active ingredient. Pharmacological effects of the compound expressed by the formula are tested on the basis of the Ames method using Salmonella typhimurium (i.e. whether reverse mutation is inhibited by a specific mutagen or not) and it is confirmed that the compound expressed by the formula exhibits powerful mutagenicity inhibitory effects. The compound expressed by the formula is obtained by reacting a trihalogermylpropionic acid with hydrogen sulfide.

Description

[Detailed Description of the Invention] [Field of Industrial Application] The present invention relates to a mutagenicity inhibitor, and more specifically to a strong mutagenicity inhibitor containing a specific organic germanium compound as an active ingredient. be.

(Prior art) Mutation is a genetic change that occurs due to a change in the relative relationship of bases in DNA in an organism.
During replication, isomeric bases generated for some reason are incorporated and separate base relationships are formed, resulting in the appearance of DNA with incorrect information, and as this process is repeated, It is estimated that this may lead to mutations.

The above-mentioned mutations include naturally occurring mutations and induced mutations caused by some factors, and the properties of compounds that cause the latter induced mutations are particularly referred to as mutagenicity.

Taking food and luxury goods as an example, in recent years, due to the development of preservation technology and the development of distribution systems, acute toxicity such as food poisoning, which is typified by microbial contamination, has become less of a problem. On the other hand, there is increasing interest in genetic toxicity that may affect not only the human being but also their descendants.

For example, whether or not a certain compound has genetic toxicity as described above is a matter that cannot be determined until experimental animals have been bred for many generations due to its nature. For example, using improved strains that have been improved to enable highly sensitive detection of mutagenic substances, etc.
Mutagenicity testing methods using microorganisms have made great progress, and in this regard, businesses that intend to introduce new chemical substances, etc. should check whether or not they are mutagenic using the above-mentioned mutagenicity testing methods using microorganisms. The law stipulates that investigations into the toxicity of such chemical substances, including substances, are required, and that if mutagenicity is found, some kind of measure must be taken.

(Problems to be Solved by the Invention) However, the above-mentioned problem of mutagenicity cannot be easily solved with regard to foods and the like that are indispensable for human life.

In other words, foods are made up of a wide variety of ingredients, and some of these ingredients may inherently contain mutagenic substances. Therefore, there is a possibility that mutagenic compounds may be mixed in, produced, or artificially added at each of these stages.

For example, heating is a typical processing means when cooking food, but it is well known that this produces mutagenic benzo(a) and ren in the cooked food.

Therefore, considering that food is essential for survival, there is an urgent need to develop drugs that can prevent mutations from occurring and prevent genetic toxicity from being transmitted to the next generation. be.

[Means for solving problems]

The present invention has been made against the background of the above-mentioned circumstances, and has the structure of It is characterized by containing an organic germanium compound represented by the following phenyl group (respectively) as an active ingredient.

That is, the inventors of the present invention investigated the mutagenicity suppressing effects of various compounds using metabolically active forms of tryptophan-like compounds, which are a type of mutagen isolated from heated amino acids and heated foods. As a result, it was discovered that the organogermanium compound represented by the above general formula exhibits a strong mutagenicity suppressing effect, and as a result of further intensive research, the present invention was completed.

The present invention will be explained in detail below.

Germanium Ge, known as a type of metal, has traditionally been the subject of research in the fields of physics and inorganic chemistry, but recently research into its organic compounds has progressed, and the results of this research have been As a result of the active publication of ``germanium'', germanium, especially its organic compounds, has attracted attention from various technical fields.

For example, carboxyethylgermanium sesquioxide (
Although G e CH2-CHI C00H) 203 exhibits not only an extremely strong blood pressure lowering effect and an effect of reducing amyloid changes, but also has excellent physiological activities such as activating macrophages and NK cells and antitumor effects by inducing interferon, The fact that it has no toxicity or side effects is becoming well known in the pharmaceutical world, and it is also being used clinically.

Since the mutagenicity inhibitor of the present invention also has an organic germanium compound represented by the above formula 1 as an active ingredient, this compound will be explained first. The basic skeleton is germylpropionic acid bonded with atoms, and the germanium atom and sulfur atom in the basic skeleton are 2
:3.

Here, the substituents R1 to R3 represent a hydrogen atom, a so-called lower alkyl group such as a methyl group, an ethyl group, a propyl group, a butyl group, or a substituted or unsubstituted phenyl group, and the substituents R1 and R2 is bonded to the α-position of the germanium atom, and the substituent R3 is also bonded to the β-position. Therefore, the following can be exemplified as the organic germanium compound used in the activator of the present invention.

(Ge-CH2-(:H, -COOH)233----
(Ia) (Ge-CHz-CI-CQOH)2s
3@66” (I c ) (:H.

(Ge-CH-CH-COOH)2s+ ”...
' (r d)CH3 CH3 (Ge-CI-CHa-COOH)*Si ” ”
” (If)aHs (Ge-CH-CH-Cool)2S+ # # #
#(Ig)Cal1°The organic germanium compound having the above structure can be produced by various methods.

That is, for example, as shown in the reaction formula below, substituent R8
For triococyanylpropionic acid such as trichlorocyamylpropionic acid (1) to which R3 has been introduced,
It is sufficient to react with hydrogen sulfide H2S.

Reaction formula The results of instrumental analysis such as nuclear magnetic resonance absorption (NMR) spectrum and infrared absorption (IR) spectrum of the organic germanium compound obtained as described above indicate that the above compound is represented by general formula 1. This is well supported, and the mutagenicity inhibitor of the present invention contains the organic germanium compound synthesized as described above as an active ingredient.
The administration form may be oral or parenteral; however, in the case of oral administration, conventional forms such as tablets, powders, or granules can be used.

(Actions and Effects of the Invention) The pharmacological effects of the agent of the present invention obtained as described above can be determined by the Ames method using Salmonella typhimurium (B, N, Ames et al.,
Proc, Natl.

Acad, Sci., 70(3), 783-786 [1
973].

That is, the above-mentioned Sa1mone11a typhimuri
um is a bacterium that cannot synthesize histidine, which is a type of amino acid (histidine auxotrophic), and when a mutagen causes back mutation, it becomes able to synthesize histidine, so the number of bacteria that have undergone back mutation The Ames method is a method for testing the presence or absence and strength of mutagenicity by examining the mutagenicity of the present agent. The test was based on whether or not it was true.

Even if the above-mentioned mutagen-inducing substances are used as they are, some of them do not show mutagenicity when assayed using the Ames method mentioned above; on the other hand, many mutagens are metabolized in vivo. It is said that the activated form reacts with biological components and exhibits a mutagen, so the actual Ames assay requires the use of the above-mentioned specific mutagen and S-9, which is prepared from rat liver microsomes. (To be precise, NA was added to the S-9 fraction prepared from rat liver microsomes.
The specific mutagen was metabolically activated by using S-9m1x containing a cofactor such as DH in combination.

Here, S prepared from the liver microsomes of the rat mentioned above
The -9 fraction is a rat liver homogenate that has been pretreated to induce the microsomal drug-metabolizing enzyme system.
This is the supernatant portion after centrifugation at ,000 Xg, and has the effect of activating the metabolism of chemical substances.

The results of the assay using the method described above are clear from the Examples described below, and the results are shown in the examples below.
It was found that there are very few bacteria in almonella typhimurium that have undergone back mutation, and that the agent of the present invention effectively suppresses mutagenicity.

〔Example〕

Next, the present invention will be explained in detail using experimental examples and examples.

Experimental Example: Synthesis of Mugermanium Compound 25.2 g (0.3 m
ol) was dissolved in 200 ml of anhydrous benzene, 2.4 g (o, 1 mol) of anhydrous pyridine was added, and after stirring, dry hydrogen sulfide gas was passed through the solution for 60 minutes. Benzene was removed while being careful of the resulting oil, the core oil was dissolved in methanol, this was added to 300 ml of purified water, and the precipitated crystals were recrystallized from methanol to obtain 17.1 g of Compound Ia.

The physical property values of the obtained compound Ia were consistent with the values described in the literature.

Other compounds were produced according to the methods described in, for example, Japanese Patent Publication No. 59-35916 or Japanese Patent Application Laid-open No. 60-16924.

EXAMPLES Examination of Pharmacological Effects ■ Preparation of Mutagen The pharmacological effects of the present invention were examined by the Ames method as described above.

The test bacterium is Salmonella typhimuri.
um strain TA-98, and the mutagen was Try-P-2, which is a type of mutagen isolated from heated amino acids and heated foods and is shown by the structural formula below.
If this Try-P-2 is not added with the above S-9 in the Ames method, Salmonella typhimuri
Since it does not show mutagenicity against the uIIl TA-98 strain, the activated Try-P-
2 was prepared in advance, and this activated Try-P-2 was used.

That is, 0.1 ml of Try-P-2 (1 mg/1 ml DMSO) solution and 1 ml of S-9mix were mixed and heated at 37°C.
After leaving for 20 minutes, 4 ml of acetone was added and stirred vigorously.

After standing for a while, centrifugation was performed at 5,000 revolutions/min for 5 minutes. The obtained supernatant was distilled under reduced pressure to remove acetone, and sterile water was added to make 2 ml.
That's what I did.

■ Assay of mutagen suppression using active ryo-P-2Sa
1monella typhimuriumTA-98
0.1 ml of bacterial solution of the strain, 1 m of active Try-P-20
l, the present invention agent (organogermanium compound from 0 to so
o Add 0.1 ml (containing 5 Lg) to top agar (a type of medium) and extract activated Tr using the standard Ames method.
When the mutagen suppression against y-P-2 was assayed, the present agent showed that active Try-P-2 was inhibited by Salmonella.
It has been found that the number of revertant colonies generated by causing reversion mutations in S. typhimurium is very small, that is, the mutagenicity of active Try-P-2 is effectively suppressed.

A part of the results of the above experiment are shown in FIG.

Although not particularly shown, other compounds may also be used in the first step.
Almost the same effect as shown in the figure was confirmed.

■Relationship with survival rate The results of the above assay show that the present agent is more effective than Salmoneltat.
It is thought that the mutagenicity of supernatant Try-P-2 was apparently suppressed by inhibiting the growth of the yphimurium TA-98 strain. The effect of suppressing mutagenicity and the survival rate of the bacteria were investigated by the Ames method using the brain incubation method.

First, add Salmonella typhim to a small test tube.
uriumTA-98 strain bacterial solution 0.5ml, active T
1 ml of ry-P-20, 0.1 ml of the present invention agent (containing 0 to 500 g of organic germanium compound), and 2.5 ml of phosphate buffer were mixed and left at 37°C for 30 minutes. The cells were collected by centrifugation at 000 rpm and suspended in 0.5 ml of phosphate buffer.

A portion of these bacteria was diluted to 104 to 10' with phosphate buffer and layered on an agar medium containing histidine to determine the number of viable bacteria. When the number of mutant strains was investigated, it was found that the agent of the present invention
The active Try
- It was confirmed that it suppresses only the mutagenicity of P-2.

A part of the results of the above experiment are shown in FIG.

Confirmed.

Although not particularly shown in the figure, other compounds also have the first
Almost the same effect as shown in the figure was confirmed.

[Brief explanation of the drawing]

FIG. 1 is a graph showing the mutagenicity suppressing effect of the agent of the present invention;
FIG. 2 is a graph showing the relationship between the mutagenicity suppressing effect of the agent of the present invention and the survival rate of bacteria.

Claims (1)

[Claims] Formulas, ▲Mathematical formulas, chemical formulas, tables, etc.▼...(I) (In the formula, R_1 to R_3 are hydrogen atoms or the same or different lower alkyl groups such as methyl and ethyl groups. or a substituted or unsubstituted phenyl group, respectively) as an active ingredient.