JPH0566943B2 - - Google Patents

Description

[Detailed description of the invention]

B. Object of the invention Industrial application field The present invention relates to a novel β-galactosidase inhibitor, galactostatin, and more specifically, the following general formula:

【formula】 (In the formula, R is

【formula】

[Formula] represents CH ₂ or C=0. ) The present invention relates to novel β-galactosidase inhibitors galactostatins, salts thereof, and methods for producing the same. Conventional technology In 3T3 fibroblasts that have become cancerous due to RNA viruses, glucosidase activity, especially β-galactosidase activity, is increased, according to H.B. Bosmann.
[Biochem.Biophys.
Acta, 264 , 339 (1972)]. Therefore, various studies have been conducted focusing on the possibility that β-galactosidase inhibitors can be used as anticancer drugs.
of Antibiotics 28 , 1006 (1975); 32 (3), 212
(1979); and 32 (3), 217 (1979). Examples of galactostatin structurally similar substances of the present invention include, for example, Japanese Patent Publication No. 43-760;
51-151393; described in JP-A-51-151394, etc., but the structure and physiological activity are different. Problems to be Solved by the Invention Examples of β-galactosidase inhibitors include those previously discovered by the present inventors and described in JP-A-57-74090.
GT-2558-A and GT-2558-B are known, but these substances are very unstable and very difficult to handle. This time, we succeeded in discovering a new stable enzyme inhibitor that is different from previously discovered β-galactosidase inhibitors. B. Means for solving the structural problems of the invention Streptomyces leideritschus PA-, the bacterium that produces GT-2558-A and GT-2558-B.
As a result of a detailed study of 5726 and 5657, in addition to GT-2558-A and GT-2558-B, β-
We discovered the existence of a weakly basic sugar analogue, 5-amino-5-deoxy-D-galactopyranose, which has galactosidase inhibitory activity and named it galactostatin. After that, we investigated the culture conditions of this strain and succeeded in improving productivity.
Galactostatin was isolated and purified in large quantities. Furthermore, the lactam and deoxy forms of galactostatin were prepared. All of these compounds have β
-It is an enzyme inhibitor that has a strong inhibitory effect on galactosidase. The properties of galactostatin, galactostatin sulfite adduct, galactostatin lactam, and 1-deoxygalactostatin are described below. Note that the present invention includes not only these compounds but also pharmaceutically acceptable salts of these compounds when possible. a Galactostatin 1 structural formula

[Formula] 2 Molecular formula/molecular weight C ₆ H ₁₃ NO ₅ MW.179.2 3 Elemental analysis value C ₆ H ₁₃ NO ₅・1/2H ₂ O Calculated value (%) C, 38, 30; H, 7.50; N7.44 Experimental value (%) C, 38.26; H, 7.47; N, 7.51 4 Melting point and decomposition temperature Melting point; 94-98°C Decomposition temperature; 113-116°C 5 Mass spectrometry spectrum (see Figure 1) Maximum mass m/z = 161 (M-18) 6 Specific rotation [α] _D ²² +85.6° (c=1.1, water) 7 Ultraviolet absorption spectrum No specific absorption 8 Infrared absorption spectrum (see Figure 2) ν _nax ^KBr 3360, 2900 , 1650, 1450, 1400, 1340,
1220, 1085, 970, 876, 806 cm ^-1 9 ¹ H nuclear magnetic resonance spectrum As shown in Figure 3, proton signals are shown at δ4.6 to 2.7 ppm. 10 Solubility Easily soluble in water, soluble in methanol, dimethyl sulfoxide, pyridine, and acetic acid. Slightly soluble in ethanol, propanol, and acetonitrile. Insoluble in acetone, ether, chloroform11 Color reaction Ninhydrin reaction Positive Ammoniacal silver nitrate reaction Positive12 Silica gel thin layer chromatography Acetonitrile:acetic acid:water (5:1:2) Rf=0.39 n-butanol:acetic acid:water (3 :1:1) Rf=0.29 13 β-galactosidase inhibition specific activity 71200IU/mg (IC ₅₀ 0.0035mcg/ml) 14 Appearance Slightly hygroscopic white powder b Galactostatin sulfite adduct 1 Structural formula

[Formula] 2 Molecular formula/molecular weight C ₆ H ₁₃ NO ₇ S・H ₂ O MW.261.3 3 Elemental analysis value C ₆ H ₁₃ NO ₇ S・H ₂ O Calculated value (%) C, 27.59; H, 5.79; N5 .36; S, 12.27 Experimental value (%) C, 27.59; H, 5.57; N5.38; S, 12.10 4 Melting point and decomposition temperature Melting point; (See Figure 4) Maximum mass m/z = 161 (M-100: -H ₂ O, -H ₂ SO ₃ ) 6 Specific optical rotation [α] _D ²³ +17.2° (c = 0.51, water) 7 Ultraviolet absorption Spectrum No specific absorption 8 Infrared absorption spectrum (see Figure 5) ν _nax ^KBr 3440, 3390, 3240, 2980, 2920, 2800,
2670, 2500, 1596, 1490, 1450, 1400, 1370,
1330, 1290, 1275, 1252, 1226, 1203, 1180,
1140, 1100, 1047, 1020, 955, 930, 805, 720,
622, 580, 525, 507, 445, 412, 357, 326, 310
cm ^-1 9 ¹ H nuclear magnetic resonance spectrum As shown in Figure 6, δ3.52, 3.50, 3.41, 3.20, 3.03,
1, 1, 1, 2, 1, and 1 protons are shown at 2.83 ppm, respectively. 10 ¹³ C nuclear magnetic resonance spectrum As shown in Figure 7, δ74.0, 72.3, 68.4, 67.6, 60.4,
^13C signal is shown at 59.9ppm. 11 Solubility Soluble in water, methanol, dimethyl sulfoxide Slightly soluble in ethanol, insoluble in acetone, ether, chloroform12 Color reaction Ninhydrin reaction Positive Ammoniacal silver nitrate reaction Positive13 Silica gel thin layer chromatography Acetonitrile:acetic acid:water) ( 5:1:2) Rf=0.55 Chloroform:methanol:ammonia (1:
2:1) Rf=0.54 14 β-galactosidase inhibition specific activity 51900IU/mg (IC ₅₀ 0.0048cmg/ml) 14 Appearance Colorless needle-like crystals c Galactostatin lactam 1 Structural formula

[Formula] 2 Molecular formula/molecular weight C ₆ H ₁₁ NO ₅ MW.177.2 3 Elemental analysis value C ₆ H ₁₁ NO ₅ Calculated value (%) C, 40.68; H, 6.26; N7.91 Experimental value (%) C, 40.40 ; H, 6.07; N7.88 4 Melting point 204-206℃ 5 Mass spectrometry spectrum (see Figure 8) Maximum mass m/z=178 (M+1) 6 Specific optical rotation [α] _D ²³ +122.0° (c= 1.0, water) 7 Ultraviolet absorption spectrum No specific absorption 8 Infrared absorption spectrum (see Figure 9) ν _nax ^KBr 3380, 3320, 2970, 2950, 2910, 1660,
1632, 1500, 1470, 1420, 1385, 1368, 1340,
1300, 1255, 1160, 1146, 1120, 1100, 1080,
1060, 1018, 985, 935, 905, 880, 830, 790,
675, 640, 600, 550, 500, 440, 370 cm ^-1 9 ¹ H nuclear magnetic resonance spectrum δ3.51, 3.49, 3.22, 3.15 ~ as shown in Figure 10
1, 1, 1, 3 protons are shown at 2.85ppm. 10 ¹³ C nuclear magnetic resonance spectrum As shown in Figure 11, δ174.3, 73.0, 70.0, 68.6,
¹³ C signals are shown at 61.7 and 55.6 ppm. 11 Solubility Soluble in water and methanol, slightly soluble in ethanol and propanol, insoluble in acetone, ether and chloroform12 Color reaction Ninhydrin reaction Positive ammoniacal silver nitrate reaction Coagulation positive13 Silica gel thin layer chromatography Acetonitrile:acetic acid:water) (5:1:2) Rf=0.61 n-butanol:acetic acid:water (3:1:1) Rf=0.34 14 β-galactosidase inhibition specific activity 43.5IU/mg (IC ₅₀ 5.75mcg/ml) 15 Appearance Colorless needle crystalline d 1-deoxygalactostatin 1 structural formula

[Formula] 2 Molecular formula/molecular weight C ₆ H ₁₃ NO ₄ MW.163.2 3 Elemental analysis value C ₆ H ₁₃ NO ₄・1/10H ₂ O Calculated value (%) C, 43.68; H, 8.06; N8.49 Experimental value (%) C, 43.68; H, 7.93; N8.31 4 Decomposition temperature 218-220℃ 5 Mass spectrometry spectrum (see Figure 12) Maximum mass m/z = 164 (M+1) 6 Specific rotation [α] _D ²³ +52.8° (c=1.0, water) 7 Ultraviolet absorption spectrum No specific absorption 8 Infrared absorption spectrum (see Figure 13) ν _nax ^KBr 3360, 2900, 1640, 1450, 1400, 1360,
1290, 1235, 1150, 1055, 935, 902, 854, 810,
715, 683, 590, 505, 475, 418 cm ^-1 9 ¹ H nuclear magnetic resonance spectrum As shown in Figure 14, δ3.33, 3.08, 2.98, 2.91,
One proton is shown at 2.80, 2.45, 2.07, and 1.70 ppm. 10 ¹³ C nuclear magnetic resonance spectrum As shown in Figure 15, δ75.9, 70.1, 69.0, 62.2,
¹³ C signals are shown at 59.7 and 49.9 ppm. 11 Solubility Easily soluble in water and methanol, soluble in ethanol and propanol, insoluble in acetone, chloroform and ether12 Color reaction Ninhydrin reaction Positive Ammoniacal silver nitrate reaction Coagulation positive13 Silica gel thin layer chromatography Acetonitrile: Acetic acid: Water ( 5:1:2) Rf=0.40 n=butanol:acetic acid:water (3:1:2) Rf=0.36 chloroform:methanol:ammonia (1:
2:1) Rf=0.49 14 β-galactosidase inhibitory specific activity 2170IU/mg (IC ₅₀ 0.115mcg/ml) 15 Appearance: Strongly hygroscopic white powder β-galactosidase inhibitory activity and inhibitory specific activity shown in the above properties are o -Nitrophenyl β-
The measurement was carried out by allowing β-galactosidase to act on D-galactopyranose as a substrate, and quantifying o-nitrophenol, which is hydrolyzed and liberated, by a colorimetric method. i.e. 0.1M acetate buffer (PH
β-galactosidase solution dissolved in 5.0) 0.25
ml, add 0.25 ml of the solution containing the specimen, warm it at 30°C for 10 minutes, and then add the substrate 0.01 Mo-nitrophenyl/β-D-galactopyranose solution dissolved in the above buffer solution previously warmed to 30°C. Mix with 0.5ml. After reacting at 30°C for 15 minutes, 1 ml of 1M sodium carbonate solution was added to stop the reaction, 8 ml of water was added to make the total volume 10 ml, and the absorbance (A) at 420 nm was measured. At the same time, the absorbance of the control using only the buffer solution without the analyte (B)
was measured, and the inhibition rate was calculated by (B-A)/B×100. The β-galactosidase used was prepared from Takadiastase. β-galactosidase activity was inhibited by 50% under the above conditions (IC ₅₀ )
The amount of the sample to be tested was defined as 1 inhibitory unit (1 Iu). Galactostatin has properties different from known β-galactosidase inhibitors, and is a novel β-galactosidase inhibitor.
- Determined to be a galactosidase inhibitor. An example of such galactostatin-producing bacteria is strain PA-5726, which was isolated from a soil sample collected from Nakadori Island, Minamimatsuura-gun, Nagasaki Prefecture. This strain was determined to be a strain of the taxonomic species Streptomyces lydicus, and strain PA-5726 was sent to the Institute of Microbial Technology as Streptomyces lydicus.
PA-5726, Microtechnical Research Institute No. 61
No. 5657). The mycological properties of the strain are as follows. (a) Morphological properties (Bennett agar medium, 28℃,
(Cultured for 14 days) No sporangia, flagellated spores, or sclerotia were observed, and no division due to fragmentation was observed in the basal hyphae. Aerial hyphae are abundantly formed on the same medium. Spore-bearing hyphae grow on aerial hyphae, and simply branch from the main axis to become side branches.
Its end exhibits a spiral shape. The surface structure of the spores observed by electron microscopy is smooth type. This strain grows over time. Part of the surface of the colony becomes wet, becoming hygroscopic and turning black. (b) Various culturing properties (cultivated at 28°C for 14 days) Color names are based on the "Color Standard" (Japan Color Research Institute version).

[Table] * The surface of the colony becomes moist, exhibits a so-called hygroscopic appearance, and turns black. Growth temperature (Bennet's agar medium, cultured at each temperature for 2 weeks. Growth at 10°C was also determined after 3 weeks.) 10°C: Judgment was made after 2 weeks, almost no growth. Judging after 3 weeks, growth is good and formation of aerial mycelia is also quite good. 28℃: Growth, aerial mycelium formation, and spore formation are all good. 37℃: No growth. 45℃: No growth. (c) Physiological properties (cultured at 28℃ for 14 days) Liquefaction ability of gelatin Positive Melanoid pigment production ability Negative Tyrosinase reaction Negative Peptonization of milk Positive Milk coagulation Positive Ability to hydrolyze starch Positive (d) Ability to utilize carbon sources Sugars commonly used for growth: L-arabinose, D-xylose, D-glucose, D-fructose, sucrose, inositol, raffinose, and D-mannite.Saccharides that showed slight growth: L-rhamnose Due to the above properties, this strain It is clear that the strain belongs to the genus Streptomyces. The Actinomycetes by Watzman
Actinomycetes) Volume 2 (1961), Shearling and Gottlieb's ISP (International
Steptomyces Project) report International Journal of Systematic Bacteriology (International Journal of
Systematic Bacteriology) Volume 18, Page 69, 279
Page (1968), Vol. 19, p. 391 (1969), No. 22
Vol. 265 (1972) and Bergey's Manual of Determinative Bacteriology.
Bacteriology) 8th edition (1974) and other publications on new species of actinomycetes PA-5726
When searching for closely related species of the strain, Streptomyces
Streptomyces lydicus
[International Journal of Systematic
Bacteriology Vol. 19, p. 448 (1969),
Bergey's Manual of Determinative
Bacteriology 8th edition, pages 772 and 777 (1974) and U.S. Patent No. 3,160,560 (1964)
year)] is listed as the most closely related species. PA-
As a result of comparing the properties of strain 5726 and those of Streptomyces lydicus described in these literatures, the properties of the two were found to match extremely well. Accordingly
The PA-5726 strain was identified as a strain belonging to the same species as Streptomyces leideritschus, and the PA-5726 strain
−5726 strain is Streptomyces leideritschus
PA−5726 (Streptomyces lydicus PA−5726)
It was named. In the present invention, not only the above-mentioned strain PA-5726 and its natural and artificial mutant strains, but also all strains that belong to the genus Streptomyces and produce galactostatin can be used.
It is within the scope of the present invention. Galactostatin production is carried out by culturing a galactostatin-producing strain in a nutrient medium under aerobic conditions, and after completion of the culturing, galactostatin is separated and collected from the culture. A general method for producing galactostatin is described below. As for the culture medium composition, culture conditions, etc., those generally used in the production of antibiotics may be used. In principle, the medium contains a carbon source, a nitrogen source, inorganic salts, etc. Vitamins, precursors, etc. may be added as necessary. Examples of carbon sources include glucose, starch, dextrin, glycerin, molasses,
Organic acids and the like can be used alone or in mixtures. Examples of nitrogen sources include soy flour, corn steep liquor, meat extract, yeast extract, cottonseed flour,
Bebutone, wheat germ, ammonium sulfate, ammonium nitrate, etc. are used alone or in a mixture. Examples of inorganic salts include calcium carbonate, sodium chloride, potassium chloride, magnesium sulfate, cobalt chloride, and various phosphates, which are added to the medium as necessary. Cultivation may be carried out according to methods generally used for the production of antibiotics, and preferably liquid culture is used.
For mass production, deep aeration culture is recommended.
If the pH of the medium is variable, a buffer such as calcium carbonate may be added to the medium. Culture is approximately 20
It is preferable to carry out the reaction at a temperature of -40°C, particularly preferably 25-32°C. Although the culture time largely depends on the scale of fermentation, the culture time required for mass production is about 20 to 80 hours. If severe foaming occurs during culturing, an antifoaming agent such as vegetable oil, lard, or polypropylene glycol may be appropriately added before or during culturing. To separate and collect galactostatin from the culture after completion of the culture, a conventional method for separating and collecting fermentation products is appropriately used. For example, filtration, centrifugation, adsorption/desorption using various ion exchange resins or other active adsorbents, chromatography, extraction using various organic solvents, etc. may be appropriately combined. It may be desirable to form galactostatin into a salt for separation operations and for convenience of use as a medicine or veterinary drug. Acids that can form salts with galactostatin include inorganic acids such as sulfite, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, phosphoric acid, and carbonic acid, as well as acetic acid, fumaric acid, malic acid, tartaric acid, and maleic acid. organic acids such as citric acid, mandelic acid, ascorbic acid, and gallic acid. Furthermore, it is possible to prepare various derivatives from the obtained galactostatin. For example, galactostatin lactam can be obtained by reacting galactostatin with an oxidizing agent such as sodium hypoiodite solution, iodine potassium iodide solution, hydrogen peroxide, ozone, or lead tetraacetate. Also,
1-Deoxygalactostatin can be prepared by catalytic reduction of galactostatin in a hydrogen stream using a catalyst such as platinum dioxide or platinum palladium, or by reacting with a reducing agent such as sodium borohydride. In addition to the above two examples, it is also possible to prepare other derivatives. These derivatives can also form salts with the acids mentioned above which can form salts with galactostatin. Examples Production examples of a group of compounds of galactostatin according to the present invention are shown below, but these examples are not intended to limit the present invention in any way. Example 1 Method for preparing galactostatin A medium (PH7.0) containing 1.0% glucose, 0.2% casamino acids, 0.2% yeast extract, and 0.1% meat extract.
Dispense 100ml each into a 500ml shake flask,
Sterilize at 120℃ for 20 minutes and use as a seed medium. Streptomyces leideritschus PA−
From the slant culture of Streptomyces lydicus PA-5726 (Streptomyces lydicus PA-5726, No. 61 (No. 5657)), a loopful of platinum was inoculated into the above seed medium.
A seed mother is prepared by culturing with shaking at 140 cycles per minute and a shaking width of 7 cm at ℃ for 2 days. On the other hand, glycerin 3.0%, meat extract 2.0%, peptone 10%, potassium chloride 0.076%, magnesium chloride hexahydrate 0.042%, iron chloride hexahydrate
Medium containing 0.0025%, zinc sulfate/heptahydrate 0.0022%, manganese chloride/tetrahydrate 0.0018% (PH7.0)
Adjust. Dispense 100 ml of this medium into a 500 ml shaking flask and sterilize it at 120°C for 30 minutes as the main fermentation medium. Inoculate 2% of the above seed mother into this main fermentation medium at 28°C at 140 cycles per minute. , perform shaking culture for 5 days at a shaking width of 7 cm. In this culture solution, the β-galactosidase inhibitor galactostatin was added at 43,500 IU/ml (IC ₅₀ =
0.0057 mcl/ml) is accumulated. Example 2 Culture was carried out in the same manner as in Example 1 using main fermentation medium 20, and immediately after the culture was completed, bacterial cells were separated by centrifugation to obtain a clear filtrate 15.8. Hydrochloric acid is added to this filtrate to adjust the pH to 4.5, 1.5% (237 g) of activated carbon is added and stirred for 30 minutes to adsorb the pigment and impurities, and filtered using Celite as an aid to obtain clear filtrate 15. This filtrate is passed through a strongly basic anion exchange resin Dowex-1×8 (OH type) (trade name, Dow Chemical Company) 2 to remove anionic substances.
This solution was neutralized with hydrochloric acid, and then passed through a column packed with 1 strongly acidic cation exchange resin DOWEX-50W x 8 (H type) to adsorb galactostatin.After thoroughly washing the resin with water, 0.5N Elute with hydrochloric acid. The β-galactosidase inhibitory activity fraction of the eluate is fractionated and collected to obtain a 1.5 galactostatin solution. The β-galactosidase inhibitory activity of this solution is
It is 238100 IU/ml (IC ₅₀ =0.0010 mcl/ml). Example 3 The galactostatin solution obtained in Example 2 is concentrated to 1/5 volume (300 ml) under reduced pressure. This concentrate contains 2
Double the volume (600 ml) of ethanol is added and the precipitated insoluble matter is removed by filtration. The obtained filtrate is further concentrated under reduced pressure to 100 ml. This solution is immediately poured into a column packed with weakly basic anion exchange resin Amberlite IRA-47 (OH type) (trade name, Rohm & Haas) 1, and the column is developed and eluted with water. The β-galactosidase inhibitory activity fraction of the eluate is fractionated and collected to obtain a galactostatin solution of 1.1. This solution is again concentrated under reduced pressure to 150 ml. To this concentrated solution, 70 ml of 6% sulfite water (Wako Pure Chemical Industries) was added under ice-cooling, and 500 ml of enotal was further added to precipitate the sulfite adduct of galactostatin as needle-shaped crystals (5.54 g). This crystal at 60℃
Dissolve in 100 ml of hot water and recrystallize by adding 400 ml of ethanol to obtain 4.85 g of colorless needle-like crystals of galactostatin sulfite adduct. β- of this crystal
Galactosidase inhibitory activity is 51900IU/mg (IC ₅₀
= 0.0048mcg/ml). Example 4 1 g of galactostatin sulfite adduct was suspended in 20 ml of water, and then a strong salt anion exchange resin Dowex-2x8 (OH type) (trade name: The Dow Chemical Company) was added.
Add 10ml and stir. The entire amount of this solution was added to the same resin dower as above.
Pour into a column packed with 200ml of 2x8 (OH type) and develop and elute with water. The β-galactosidase inhibitory activity fraction of the eluate was fractionated and concentrated under reduced pressure to obtain concentrated solution 2.
Add 20 ml of ethanol per ml to further concentrate to form a white precipitate. This precipitate is filtered and dried to obtain a white powder of galactostatin free base.
You get 628 mg. The β-galactosidase inhibitory activity of this powder is 71200IU/mg (IC ₅₀ = 0.0035mcg/ml)
It is. Example 5 Method for preparing galactostatin lactam 200 mg of galactostatin is dissolved in 5 ml of water, and while stirring, 2 ml of 0.2N iodine solution containing 4% potassium iodide is added and reacted for 5 minutes. to the reaction solution
After adding 3 ml of 0.2N sodium hydroxide solution and stirring for 5 minutes, add 2 ml of iodine solution again and repeat the above operation 6 times. After adding 12 ml of iodine solution and 18 ml of sodium hydroxide solution, add another 50 minutes. Stir and carry out the reaction. The entire amount of this reaction solution was poured into a column packed with 20 ml of a strongly acidic cation exchange resin Dowex-50W x 8 (H type) and developed and eluted with 30 ml of water. 2 g of silver carbonate is added to this eluate while stirring, and after 30 minutes, the formed precipitate is removed by filtration to obtain a clear filtrate. This filtrate was further poured into a column packed with 10 ml of a strongly acidic cation exchange resin Dowex-50W x 8 (H type) and developed and eluted with 20 ml of water. This eluate is neutralized by adding weakly basic anion exchange resin Amberlite IRA-47 (OH type) to reduce its slightly acidic nature.
The resin is removed by filtration to obtain a clear, neutral filtrate. The filtrate was concentrated under reduced pressure and ethanol was added to produce galactostatin lactam as needle crystals (105
mg). Dissolve these crystals in 1 ml of water, add 15 ml of ethanol and recrystallize to obtain colorless needle-shaped galactostatin lactam crystals.
You get 95.1 mg. The β-galactosidase inhibitory activity of this crystal is 43.5 IU/mg (IC ₅₀ =5.75 mcg/ml). Example 6 Method for preparing 1-deoxygalactostatin 200 mg of galactostatin and 5 ml of 50% methanol
40 mg of platinum dioxide and 0.2 ml of acetic acid were added, and catalytic reduction was carried out for 5 hours in a hydrogen stream under normal pressure while stirring. After the reaction is completed, the supernatant obtained by centrifugation is filtered to obtain a clear filtrate. After concentrating this filtrate under reduced pressure, 25 ml of strongly basic anion exchange resin Dowex-2×8 (OH type)
Pour into a column packed with water, develop and elute with water. The β-galactosidase inhibitory activity fraction of the eluate is fractionated and concentrated under reduced pressure to obtain an oily substance. This oil is dissolved in 1 ml of menothal and 2 ml of acetone and 10 ml of ether are added to form a white precipitate. This precipitate is collected by filtration and dried to obtain 107 mg of white powder of 1-deoxygalactostatin. The β-galactosidase inhibitory activity of this powder is
It is 2170IU/mg (IU ₅₀ =0.115mcg/ml). Effects of the invention Galactostatin is very stable in acidic and neutral solutions, hardly undergoing any decomposition changes, and is highly stable in acidic and neutral solutions.
2558 is also relatively stable in acidic and neutral solutions. However, in a basic solution, GT-2558 rapidly browns and decomposes, and is extremely unstable to the extent that silica gel thin layer chromatography shows an increase in tailing substances from a different origin from that of GT-2558. For statins, only slight tailing was observed. As long as galactostatin powder is stored at 4°C in a cool place, it does not change color or decrease in β-galactosidase inhibitory activity, and is stable for a long period of time, whereas GT-2558 has a slight yellow coloration. As a result, the β-galactosidase inhibitory activity also gradually decreases. Furthermore, the difference in stability between galactostatins and GT-2558 is also reflected in the difference in recovery rate from the culture filtrate of actinomycetes. The recovery rates of galactostatin and GT-2558 are shown below.

【table】

[Table] Galactostatin, its constituent components, and their pharmaceutically acceptable salts can be used as galactosidase inhibitors and reagents for measuring galactosidase activity. Furthermore, as mentioned above, they have the possibility of being used as anticancer agents, in which case these compounds are administered orally or parenterally to humans or animals. Using commonly used excipients, stabilizers, preservatives, wetting agents, surfactants, etc., it can be administered orally in the form of tablets, capsules, or powders, or as injections, liniments, suppositories, etc. It can also be administered parenterally, and the dosage varies depending on the therapeutic purpose, age of the patient, symptoms, etc. Furthermore, the compounds of the present invention have antibacterial and antiviral activities, although they are weak.

[Brief explanation of drawings]

FIG. 1 is a mass spectrometry spectrum of galactostatin, and FIG. 2 is an infrared absorption spectrum of galactostatin in potassium bromide tablets. FIG. 3 is a 200MHz ¹ H nuclear magnetic resonance spectrum of galactostatin, measured in 0.5N dihydrochloric acid in heavy water using tetramethylsilane as a standard substance. FIG. 4 is a mass spectrometry spectrum of galactostatin sulfite adduct, and FIG. 5 is an infrared absorption spectrum of galactostatin sulfite adduct in potassium bromide tablets. Figures 6 and 7 are nuclear magnetic resonance spectra of galactostatin sulfite adduct, Figure 6 is a 200MHz ¹ H nuclear magnetic resonance spectrum measured using tetramethylsilane in heavy water as a standard substance, and Figure 7 is a nuclear magnetic resonance spectrum of galactostatin sulfite adduct. This is a 25MHz ¹³ C nuclear magnetic resonance spectrum measured using acetonitrile in heavy water as a standard substance. FIG. 8 is a mass spectrometry spectrum of galactostatin lactam, and FIG. 9 is an infrared absorption spectrum of galactostatin lactam in potassium bromide tablets. Figure 10 and 1
Figure 1 is a nuclear magnetic resonance spectrum of galactostatin lactam, Figure 10 is a 200MHz ¹ H nuclear magnetic resonance spectrum measured in heavy water using tetramethylsilane as a standard substance, and Figure 11 is a nuclear magnetic resonance spectrum measured in heavy water with acetonitrile as a standard substance. 25MHz measured as ¹³
C nuclear magnetic resonance spectrum. Figure 12 is 1-
This is a gravimetric analysis spectrum of deoxygalactostatin, and FIG. 13 is an infrared absorption spectrum of 1-deoxygalactostatin in potassium bromide tablets. Figures 14 and 15 are nuclear magnetic resonance spectra of 1-deoxygalactostatin,
FIG. 14 shows a 200 MHz ¹ H nuclear magnetic resonance spectrum measured using tetramethylsilane in heavy water as a standard substance, and FIG. 15 shows a 25 MHz ¹³ C nuclear magnetic resonance spectrum measured in heavy water using acetonitrile as a standard substance.

Claims (1)

[Claims] 1 A compound represented by the formula: [Formula] (wherein R represents [Formula] [Formula] CH ₂ or C = 0) or a salt thereof. 2. A method for producing galactostatin, which comprises culturing galactostatin-producing bacteria belonging to the genus Streptomyces in a medium, and separating and collecting galactostatin from the culture.