JPS5924134B2 - Antihypertensive agents containing aminobenzoic acid derivatives - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an antihypertensive agent containing a substance represented by general formula (1) or a salt thereof as a main component.

R-NHq (1) (where R1 represents arabinose, xylose, glucose, galactose, rhamnose residue) Conventionally, synthetic compounds and antibiotics have been used as anticancer agents, but these have excellent cancer killing effects. However, it has the disadvantage of being highly toxic and causing side effects because it also acts on normal cells.

Therefore, recently, polysaccharides of various origins have attracted attention because they exhibit anticancer effects by enhancing the host's immune function. The present inventors have already developed and provided to society an anticancer agent made from a polysaccharide derived from basidiomycetes, but during research into the structure and activity of this anticancer agent, we discovered that aminobenzoic acid-N-D-mannoside had an antitumor effect, We found a lowering effect and a blood pressure lowering effect. Subsequently, as a result of further research and improvement, a compound represented by the above general formula (1) with low toxicity and higher medicinal efficacy was discovered, and the present invention was completed. General formula (1
) (hereinafter abbreviated as Substance 2) has a simple structure, but has extremely low toxicity and no antibacterial activity, so it can be administered for a long period of time without worrying about disrupting the intestinal flora. It is.

Furthermore, it is an extremely safe drug that does not have mutagenicity or affect cell-mediated or humoral immunity, and therefore poses no risk of teratogenicity or allergic reactions in healthy people. In addition, all of these substances have a blood pressure lowering effect and are useful as antihypertensive agents. There are three types of amino group positions relative to the carboxyl group of this substance: p-, m-, and o-.
Although there may be some differences in activity, all are essentially useful. In addition, the salt of the aminobenzoic acid derivative of the present invention is one in which the hydrogen atom of the -COOH group in the above formula (1) is replaced with an alkali metal, an alkaline earth metal, or an aluminum metal. These metals may be any metal as long as it is acceptable as a drug, and are usually Na, . K. MglCa,. Al is preferable, and Na is particularly preferable. In addition, the sugar part is 6
It has a membered ring (pyranose) structure. The manufacturing method of this substance is as follows.

Aminobenzoic acid 4.5-57, sugar (L-arabinose,
D-xylose, D-glucose, D-galactose or L-rhamnose) 5-67, ammonium chloride 0.
1 to 0.5y was heated and condensed under reflux of 40 to 90 ml of 95 to 100% ethanol or pure methanol.

If crystals precipitated after being left at room temperature or in a cold place for a while, the reaction solution was thoroughly filtered, the crystals were thoroughly washed with water, alcohol, ether, etc., and then recrystallized from methanol water or ethanol water. A well-known method was used to replace the hydrogen of the carboxyl group with a base.

That is, this substance was dissolved in an alcoholic aqueous solvent and an inorganic salt was added for substitution. The physicochemical properties of this substance obtained by the above production method are shown in Table 1 below. Further, infrared absorption spectra are shown in Figs. 1-20. The analysis methods in Table 1 are as follows. (1) Melting point Measured using a Yanagimoto micro melting point measuring device.

(2) Optical rotation H
is an alcohol-aqueous system, and -Na is an optical path length of 5 using water as a solvent.
Measured at 0 mm.

(3) Elemental analysis Measured using Yanagimoto CHN coater MT2 type.

(4) Using UV Hitachi EPS-3T self-recording spectrophotometer,
-H was measured using an alcohol-aqueous system, and -Na was measured using water as a solvent.

(5) Measured by the KBr method using IR Japan Spectroscopy DS-701G model.

Note that this corresponds to the sample waterfall in Table 1 of the drawing.

Next, the toxicological properties of this substance are shown.

(1) Acute toxicity Acute toxicity was investigated by intraperitoneal and forced oral administration using ICR-JCL mice.

This substance was dissolved in physiological saline for intraperitoneal administration, and in distilled water for oral administration, and the solution was adjusted to a predetermined amount using a syringe or a stomach tube and administered. After administration, the symptoms of toxicity were continued to be observed, and the LD5O value was determined from the mortality rate over time up to the 7th day.

The findings were obtained through autopsy in both surviving and dead cases. The LD5O value is determined by the Litchfield-Wilcoxon (Litchfie)
ld-WilcOxOn) graphic calculation method.

The results are shown in Table 2. Both are LD regardless of intraperitoneal or oral administration.
With a 5O value of 67/Kg or more, it is a low-toxic substance that falls well into the category of so-called "ordinary drugs," and 6 out of 10 compounds, more than half, have an LD5O value of 107/K9.
Based on the above, it can be said that this drug is extremely safe. (2)
Antibacterial activity This substance was dissolved in distilled water to prepare a 2-fold dilution series, and this dilution was mixed with 9 times the volume of heated and dissolved agar medium, poured into a Petri dish, and plated.

Hartin Fusion agar (bacteria) and Sabouraud agar (fungi) were used as the culture medium, and after inoculating the pre-cultured test bacteria, the bacteria were incubated at 37°C for 20-24 hours. The fungi were cultured at 25° C. for 3 to 7 days, and the presence or absence of growth was examined. The following bacterial species were used as test bacteria. Pseudomonas aeruginosa
asaeruginOsaIAMl5l4) Escherichia coli (E
scherichiacOliIFOl2734) Staphylococcus aureus
s2O9P) Bacillus subtili
sIAMlO69) Baker's Yeast (SaccharOmyc)
Candida albicans ATCC752
TrichOphytOnmentagrOp
hytesIFO6l24) Black mold (Aspergil)
As a result, this substance did not inhibit the growth of any bacteria at a concentration of 1η/ml.

(3) Mutagenicity First, an investigation using Rec-Assay was conducted.

That is, a tissue repair-deficient strain (Baeillus subt.
ilis M45) and the recombinant repair carrier strain (B. subtil
lsHl7) on B-agar medium (Nikukozu 107,
Polypeptone 107, NaCl5y, Agar 155! , distilled water 1000ml,. (pH 7.0) so that the starting points did not touch each other.

Dissolve this substance in sterilized water, absorb 0.04 ml of the solution onto a circular paper with a diameter of 8 squares, and immediately let it stand so as to cover the starting point of the streak, and culture it overnight at 37°C to inhibit growth. The length of the area was measured. Kanamycin was used as a negative control, and mitomycin C was used as a positive control. Next, perform the reverse mutation test with S
This was carried out using almOnellatyphimurium TA98 and TAlOO (both require histidine).

Soft agar solution (6y NaCl, 6y agar, 11% distilled water) containing 0.5mM biotin-0.5mM histidine solution
000ml) 2ml with 0.1ml of bacterial solution and 0.1mj of chemical solution
was added, mixed well, and layered on a minimal agar medium.

The cells were cultured at 37°C for 2 days and the number of revertant colonies was counted.

Furilfuramide (AF2) was used as a positive control. The results of the Rec-Assay are shown in Table 3, and the results of the reverse mutation test are shown in Table 4.

In the Rec-Assay, this substance did not lack mutagenicity even at high concentrations, but the sodium P-aminobenzoate derivative was particularly excellent. In addition, in the reverse mutation test, no change was observed in the mutation incidence rate due to this substance compared to the non-additive control even when a high concentration was applied, proving that it is a highly safe drug. (4) Delayed intradermal reaction In order to understand the effect of this substance on cell-mediated immunity, we used ICR-JCL mice to perform a foot-step reaction (FOOt) using sheep red blood cells as an antigen.
padreactiOn) was performed.

Sheep red blood cells were suspended in a 10% volume in physiological saline, and 0.2 d of this solution was injected through the tail vein for primary sensitization, and after 7 days, 0.05 m of a 40% suspension of sheep red blood cells was added.
1 was injected into the heel of the foot for secondary sensitization, and the foot jump thickness was measured the next day. This substance is 250η around the day of primary sensitization.
/Kg was administered intraperitoneally 5 times daily. As a result, no significant difference was observed in the increase in foot thickness in the group administered with this substance compared to the control (non-administration) group.

(5) Antibody production ability In order to understand the effect of this substance on humoral immunity, ICR-J
For CL mice, a 10% volume suspension of Higgi's red blood cells was administered at 0.
2mj was injected into the tail vein for sensitization, and on the 7th day after sensitization, blood was collected and antibody production ability was measured by hemagglutination reaction.

The substance was intraperitoneally administered at a dose of 250 K9 five times on consecutive days, mainly on the day of sensitization. As a result, no significant difference was observed in the agglutination value between the group administered with this substance and the control group.

Next, we will discuss the pharmacological properties of this substance.

1) Blood pressure lowering effect Dissolved in distilled water on spontaneously hypertensive rats (SHR) with a systolic blood pressure of 200 to 210 mmHg at 20 to 25 weeks of age, which is known as the best model animal for human instinctive hypertension. 30W/K9 or 3 of this substance
It was orally administered at a rate of 00η/K9.

3 hours after administration. and 6hr. Tail artery pressure was measured non-invasively using a blood pressure measuring device (manufactured by Ueda Seisakusho, model USM-105R). The number of animals in each group was 5. The results of the average values are shown in Table 5. All of these substances clearly exhibit antihypertensive effects and are useful as antihypertensive agents. Sodium O-aminobenzoate N-L-arabinoside, Sodium P-aminobenzoate-N-D-xyloside, P and O
-Sodium aminobenzoate-N-D-glucoside, P
The hypotensive effect of mono- and O-aminobenzoic acid-N-L-rhamnoside is as large as 20 mmHg or more, and in particular, sodium P-aminobenzoate-N-D xyloside and sodium P-aminobenzoate N-D-glucoside are both 30 mmHg or higher.ヮ／K
32-35 mTI at oral doses as small as 9, respectively.
It was known to have an excellent effect, showing a reduction in blood pressure of LHg and 23 mm 1 Ig.

In general, even if drugs are highly effective, many have strong side effects and cannot be administered over a long period of time.

As mentioned above, this *substance not only has low acute toxicity, but also has no mutagenicity, no allergenicity, and no adverse effects on intestinal flora.
The LD5O value is at least 250 times higher than that of 30 Immediate/K9, which shows medicinal efficacy, and the safety factor is extremely high. From this point of view as well, this substance can be said to be a drug that can be safely administered over a long period of time. Initiated hypertension is a long-term disease, and once the administration of antihypertensive drugs has been started, it is impossible to stop taking them.In this sense, the present substance can be said to be useful as a hypotensive drug. Next, we will discuss the formulation of this substance.

When used as an antihypertensive agent, this substance can be used in any convenient form to obtain its medicinal effect depending on the type and symptoms of the disease, and may be used alone or in a mixture with pharmaceutically acceptable diluents and other drugs. Can be used.

The substance is applied orally or parenterally.

Therefore, it may take any form for oral or parenteral administration. The substances can be provided in dosage unit form.

They contain an effective amount of the active ingredient and can take the form of powders, granules, tablets, dragees, capsules, suppositories, suspensions, solutions, emulsions, ampoules, injections, and the like. The diluent may be solid, liquid, semi-solid, or in the form of an ingestible capsule, such as the following: i.e. excipients, fillers, binders, wetting agents, disintegrants, surfactants,
These include lubricants, dispersion buffers, fragrances, preservatives, solubilizing agents, and solvents. Furthermore, one type or a mixture of one or more types of these may be used. The medicament of the present invention can be produced by any known method.

The active ingredients in the compositions used in the present invention generally range from 0.01% to 100% by weight. %included. The pharmaceutical agent of the present invention can be administered orally or parenterally to humans and animals, but oral administration is preferred. Oral administration includes sublingual administration. Parenteral administration includes injections such as subcutaneous, intramuscular, intravenous, infusion, and the like. The dosage of the medicine of the present invention depends on whether it is an animal or a human being, and is influenced by age, individual differences, medical conditions, etc., and in some cases, doses outside the range shown below may be administered.
Generally, when targeting humans, the oral dosage of this substance is 0.1 to 500 η per day, preferably 1 to 250 Tf19, and the parenteral dosage is 0.0
1 to 200η, preferably 0.1 to 100η once to 4 times
Administer in divided doses.

Hereinafter, the present invention will be explained in more detail by showing formulation examples and manufacturing examples of the substance of the present invention.

Formulation example 1 Mix them uniformly to make a powder or fine granules.

Further, this powder was put into a capsule container to form a capsule. After uniformly mixing and blending Formulation Example 2, the mixture is crushed, granulated, dried, and sieved to form granules.

Preparation: Example 3 Sodium O-aminobenzoate N-D-xyloside in Example 2 was replaced with sodium O-aminobenzoate-N
- Make granules using the same method using D-glucoside,
4 parts of calcium stearate are added to 96 parts of the granules and compression molded to form tablets with a diameter of 10 mm.

Formulation example 4 Granules are prepared in the same manner as in Example 2 using

10 parts of crystalline cellulose is added to 90 parts of the obtained granules and compression molded to form 8-fold diameter tablets, and syrup gelatin and precipitated calcium carbonate are added to make sugar-coated tablets.
Formulation Example 5 is heated and mixed and then sterilized to prepare an injection.

51. 0.5V of ammonium chloride is heated and condensed in 40ml of 194% ethyl alcohol under reflux.

If the reaction solution is left in the refrigerator, crystals will precipitate. The reaction solution was passed through the mouth, the crystals were washed with ether, and recrystallization was repeated several times from 50% methyl alcohol to obtain colorless needle-shaped crystals. The yield was 45.8%. The thus obtained P-aminobenzoic acid monoN-L-arabinoside was gradually dissolved in a 1% aqueous solution containing a calculated amount of NaOH, the insoluble matter was sifted through the mouth, and the oral fluid was concentrated under reduced pressure. Add excess acetone,
After dehydration and drying, colorless crystals were obtained. The yield was 100%, and the total yield was 45.8%. Production Example 2 Production of 0-aminobenzoic acid-N-L-arabinoside monosodium salt Anthranilic acid 2.37, L-arabinose 2.
57. 0.2 V of ammonium chloride is heated and condensed in 30 ml of methyl alcohol under reflux.

After the reaction, if left at room temperature, crystals will precipitate.

The reaction solution was passed through the mouth, and the crystals were washed with water, methyl alcohol, and ether to obtain colorless needle-like or disk-like crystals. Yield 61
．． It was 3%. The thus obtained anthranilic acid-NL-arabinoside containing a calculated amount of NaOH
% aqueous solution, insoluble matter was filtered out, the oral liquid was concentrated under reduced pressure, a large excess of acetone was added, and after dehydration, the mixture was dried to obtain colorless crystals.

The yield was 100%, and the total yield was 61.3%. Production Example 3 Production method of P-aminobenzoic acid-N-D-xyloside monosodium salt P-aminobenzoic acid 2.37, D-xylose 2.
57. 0.05y of ammonium chloride is heated and condensed in 25ml of ethyl alcohol under reflux.

Although crystals precipitate during the reaction, add more solvent and continue heating to complete the reaction.

After standing in a cold place, the reaction solution was sifted, the crystals were washed with water, dilute methyl alcohol, and a small amount of ether, and recrystallized from 94% ethyl alcohol to obtain colorless needle-shaped crystals. The yield was 73.7%. P obtained in this way
-Aminobenzoic acid N-D-xyloside in a calculated amount of NaO
Gradually dissolve in a 1% aqueous solution containing H, pass through the insoluble matter,
The oral fluid was concentrated under reduced pressure, a large excess of acetone was added, and after dehydration,
Drying gave colorless crystals.

The yield was 100%, and the TOtal yield was 73.7%. Production Example 4 Production method of 0-aminobenzoic acid-N-D-xyloside monosodium salt Anthranilic acid 2.37V, D-xylose 2.5V
0.27 ammonium monochloride is heated and condensed in 35 ml of ethyl alcohol under reflux.

After the reaction, the mixture is concentrated under reduced pressure to approximately 1%, and when it is left at room temperature, precipitation of crystals is observed.

The reaction solution was passed through the mouth, the crystals were washed with water, methyl alcohol, and ether, and recrystallized from ethyl alcohol to obtain colorless needle-like crystals. The yield was 74.6%. The thus obtained anthranilic acid-N-D-xyloside was gradually dissolved in a 1% aqueous solution containing a calculated amount of NaOH, the insoluble material was filtered off, the oral solution was concentrated under reduced pressure, and a large excess of acetone was added. was added, dehydrated and dried to obtain colorless crystals.

The yield was 100%, and the TOtal yield was 76.4%.

Production Example 5 Method for producing P-aminobenzoic acid-N-D-glucoside-Na salt P-aminobenzoic acid 5V. ．． D-glucose 6.4
f. 0.5f1 of ammonium chloride is heated and condensed in 50ml of 194% ethyl alcohol under reflux.

After the reaction, the mixture was concentrated under reduced pressure to approximately 1%, and when left in a cold place, the entire liquid became gelatinous.

Add a small amount of water, warm again to dissolve, and then leave it in the refrigerator to see crystals precipitate. Pass the reaction solution through the mouth, wash the crystals with water, dilute methyl alcohol and a small amount of ether,
Recrystallization from 0% methyl alcohol gave colorless needle-like crystals.

The yield was 33.7%. Obtained in this way, P
-aminobenzoic acid-N-D-glucoside in a calculated amount of Na
It was gradually dissolved in a 1% aqueous solution containing OH, the insoluble matter was filtered out, the oral solution was concentrated under reduced pressure, a large excess of acetone was added, and after dehydration, it was dried to obtain colorless crystals.

The yield was 100%, and the TOtal yield was 33.7%. Production Example 6 Production method of 0-aminobenzoic acid-N-D-glucoside-Na salt Anthranilic acid 4.6t,. D-glucose 6.0
y1 0.57 ammonium chloride is heated and condensed in 40 mL of 195% ethyl alcohol under reflux.

After the reaction, concentrate under reduced pressure to approx. % and leave it in the refrigerator overnight to observe the precipitation of crystals.

The reaction solution was passed through the mouth, the crystals were washed with water, methyl alcohol, and ether, and recrystallized twice from methyl alcohol to obtain colorless needle-like crystals.

The yield was 4.6%. The anthranilic acid-ND-glucoside thus obtained was gradually dissolved in a 1% aqueous solution containing a calculated amount of NaOH, the insoluble material was filtered off, the oral fluid was concentrated under reduced pressure, and a large excess of acetone was added. After dehydration and drying, colorless crystals were obtained. Yield 100%, TOtal yield 4.6
It was %. Production Example 7 Production method of P-aminobenzoic acid-N-D-galactoside Na salt P-aminobenzoic acid 1.57, D-galactose 2
y1 0.1y of ammonium chloride is heated and condensed in 30ml of 194% ethyl alcohol under reflux.

After the reaction, concentrate under reduced pressure and leave in a cool place to observe precipitation of crystals. The reaction solution was passed through the mouth, and the crystals were washed with water, diluted methyl alcohol and a small amount of ether, and recrystallized from methyl alcohol to obtain colorless needle-like crystals.

The yield was 18.1%. Obtained in this way, P
-Aminobenzoic acid mono-N-D-galactoside in the calculated amount of N
It was gradually dissolved in a 1% aqueous solution containing aOH, the insoluble matter was filtered out, the oral liquid was concentrated under reduced pressure, a large excess of acetone was added, and the mixture was dehydrated and dried to obtain colorless crystals.

The yield was 100%, and the TOtal yield was 18.1%. Production Example 8 Production method of 0-aminobenzoic acid-N-D-galactoside-Na salt Anthranilic acid 2.47, D-galactose 3.
07. 0.2y of ammonium chloride is heated and condensed in 30ml of 195% ethyl alcohol under reflux.

After the reaction, the reaction solution is concentrated under reduced pressure to approx. Needle-shaped crystals were obtained.

The yield was 16.4%. The thus obtained anthranilic acid-ND-galactoside was dissolved in a calculated amount of NaOH.
The mixture was gradually dissolved in a 1% aqueous solution containing 1% aqueous solution containing insoluble materials, filtrated to remove insoluble matter, and the oral solution was concentrated under reduced pressure. A large excess of acetone was added, dehydrated, and then dried to obtain colorless crystals.

The yield was 100%, and the TOtal yield was 16.4%.

Production Example 9 Production method of P-aminobenzoic acid-N-L-rhamnoside monosodium salt 31 P-aminobenzoic acid, 4f1 L-rhamnose, 0.17 ammonium chloride are condensed by heating in 94% ethyl alcohol under reflux cooling. .

After the reaction, if left at room temperature, crystals will precipitate.

The reaction solution was passed through the mouth, the crystals were washed with water and diluted methyl alcohol, and then recrystallized from 50% methyl alcohol to obtain colorless needle-shaped crystals.

The yield was 30.9%. Obtained in this way, P
- Aminobenzoic acid mono-N-L-rhamnoside 0.27 was gradually dissolved in a 1% aqueous solution containing the calculated amount of NaOH, the slurry was sifted through the mouth, the oral fluid was concentrated under reduced pressure, and a large excess of acetone was removed. In addition, the mixture was dehydrated and dried to obtain colorless crystals.

The yield was 100%, and the TOtal yield was 30.9%. Production Example 10 Production method of 0-aminobenzoic acid-N-L-rhamnoside monosodium salt Anthranilic acid 2.37y, L-rhamnose 2.8y
0.27 ammonium monochloride is heated and condensed in 25 ml of methyl alcohol under reflux.

After the reaction, if left at room temperature, crystals will precipitate.

The reaction solution was passed through the mouth, the crystals were washed with water and methyl alcohol, and then recrystallized from 50% methyl alcohol to obtain colorless needle-shaped crystals. The yield was 9.8%. Anthranilic acid-NL-rhamnoside 0.2 thus obtained
? was gradually dissolved in a 1% aqueous solution containing a calculated amount of NaOH, impurities were filtered out, the oral liquid was concentrated under reduced pressure, a large excess of acetone was added, and the mixture was dehydrated and dried to obtain colorless crystals.

The yield was 100%, and the TOtal yield was 9.8%.

[Brief explanation of the drawing]

1 to 20 are infrared absorption spectrum diagrams of the substances of the present invention.

Claims (1)

[Claims] 1 At least one aminobenzoic acid derivative or its salt represented by the general formula ▲ Numerical formula, chemical formula, table, etc. ▼ (where R_1 represents an arabinose, xylose, glucose, galactose, rhamnose residue) Antihypertensive agent containing seeds. 2 General formula ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼ (However, R is arabinose, xylose, glucose,
The antihypertensive agent according to claim 1, which is a galactose or rhamnose residue. 3. The antihypertensive agent according to claim 1 or 2, which is in an oral dosage form. 4. The antihypertensive agent according to claim 1 or 2, which is in a parenteral administration form.