JPH0369894B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a novel compound bethenone C useful as a physiologically active substance and a method for producing the same.

Formula of the invention Bethenone C represented by is a novel compound having the following physical and chemical properties.

(1) Shape: Oil-like (2) Specific rotation: [α] _D −38.6° (C4.15, CHCl ₃ ) (3) Molecular weight by high-resolution mass spectrum: obs 366.2418 calcd366.2406 (4) Molecular formula: C ₂₁ H ₃₄ O ₅ (5) Ultraviolet absorption spectrum: λ ^EtOH _nax 276nm (ε=5765) (6) Infrared absorption spectrum (main peak): IRν ^neat _nax cm ^-1 3470, 2950, 2870, 1700,
165, 1570, 1440, 1370, 1250, 1130, 750 (7) Thin layer chromatography [using silica gel (Merck)]: Solvent system Rf value Chloroform: methanol (95:5, v/
v) 0.65 (8) Solubility: Soluble; methanol, chloroform, ethyl acetate Insoluble: Hexane (9) Color reaction: Anisaldehyde reagent (spraying, heating), blue-green (10) Nuclear magnetic resonance spectrum: 'H- NMRδ ^CDCl3 _TMS (400MH ₂ ) 0.74 (3H, d, 6.8Hz), 0.79 (3H, t, 7.3
Hz), 1.12 (3H, d, 6.8Hz), 1.16 (1H,
dd, 12.2, 13.7Hz), 1.26 (3H, s), 1.31
(3H, s), 1.33-1.50 (2H, m), 1.55
(1H, s), 1.61 (3H, s), 1.74-1.92
(2H, m), 2.33 (1H, dt, 3.4Hz, 13.7Hz),
2.58 (1H, dd, 9.8Hz, 12.7Hz), 2.65 (1H,
dt, 3.9Hz, 12.7PH), 5.89 (1H, d, 4.9
Hz), 7.74 (1H, d, 4.9Hz), 15.13 (1H,
br, s) The biological properties of the present compound Bethenone C are as follows. That is, the inhibition of DNA, RNA, and protein synthesis by starfish gastrulae by this compound is as shown in the figure, and the measurement conditions are as follows: [ ³ H] thymidine and [ ³ H] as precursor substances in 0.5 ml of physiological saline containing 3200 gastrulae. After adding 0.2 to 0.4 μCi of uridine or [ ³ H]leucine and reacting at 20° C. for 1 hour, the incorporation rate of the precursor substance into the acid-insoluble fraction was determined. As is clear from FIG. 1, this compound suppresses the uptake of each precursor substance and inhibits DNA, RNA, and protein synthesis.

As the strain used to produce the compound bethenone C of the present invention, a betenone C-producing bacterium belonging to Homa bethe is used. For example, Homa Bethe Frank (The British
Mycological Society Transactions Vol.46,
Part 4, 1963, page 614)
Bethe Frank PS-13 (Phoma betae Frank
PS-13) has been deposited with the Institute of Microbial Technology, Agency of Industrial Science and Technology as Microbiological Research Institute No. 6556.

The bacteria to be used in the present invention include not only the above-mentioned Homa-Bethe-Frank PS-13 and mutant strains of this strain, but also any bacteria that produce the compound bethenone C newly isolated from nature. can.

In the present invention, the carbon source and nitrogen source used in the culture medium are those available for the strain used. Examples of the carbon source include starch, glycerin, dextrin, sucrose, maltose, glucose, and potato starch. Nitrogen sources include peptone, meat extract, yeast extract, casein hydrolyzate, corn steep liquor, gluten meal, corn meal, soybean flour, wheat germ, cottonseed meal, ammonium sulfate, ammonium nitrate, Urea etc. are used. In addition, salts such as phosphate, magnesium, potassium, calcium, sodium, iron, and manganese are used as necessary.

Cultivation is carried out under aerobic conditions such as static culture or aerated agitation culture. The culture temperature is usually 20 to 35°C, and the culture is terminated at an appropriate time by determining the time when bethenone C reaches the maximum production concentration.

After completion of the culture, bethenone C is collected from the culture. For example, by separating the culture into bacterial cells and liquid,
After extraction from the liquid with an organic solvent (for example, ethyl acetate) that dissolves Bethenone C, Bethenone C is recovered by a known method commonly used in the purification of fat-soluble substances. For example, after concentrating the extract, bethenone C is isolated by silica gel column chromatography.

Next, examples of the present invention will be shown, but these are merely examples and do not limit the present invention.

Example: Cut 1,000 g of peeled yam into 1 cm cubes, add about 5 ounces of tap water, and place in an autoclave (1 cm).
Kg/cm ² for 10 minutes) and pass the decoction through 4 pieces of gauze to obtain about 5. 20g sugar for this
was added and further heated in an autoclave for 5 minutes to dissolve the medium, which was then dispensed into 150 ml portions into 500 ml Erlenmeyer flasks and sterilized. In this, Homa Bethe Frank
PS-13 (Feikoken Bacteria No. 6556) was inoculated and statically cultured at 25°C for 15 days. Next, all the cultures containing Bethenone C were collected and the bacterial cells were removed by excessive removal to obtain about 20 liquids. This was concentrated under reduced pressure to a pH of 5.2 at a temperature of 40° C. or below, glacial acetic acid was added to adjust the pH to 5.5, and 3.5 of ethyl acetate was added for extraction.
This extraction operation was repeated five more times, and the resulting extract was combined with the first extract and concentrated under reduced pressure to obtain about 5.6 g of a dark brown oily substance. Next, this was placed on a silica gel column (Wako Gel C-200, manufactured by Wako Pure Chemical Industries, Ltd., 450 g), and the mixture was diluted with chloroform:methanol (98:2, v/v) to approximately 1.8 g.
After elution, the eluate containing Bethenone C is about 130
Got ml. A portion of the eluate was taken and subjected to silica gel thin layer chromatography (Merck, silica gel).
60F ₂₅₄ , 0.2 mm, developing solvent chloroform:methanol = 95:5, v/v, coloring agent anisaldehyde reagent), and collected fractions containing a substance exhibiting an Rf value of 0.65, and concentrated to dryness under reduced pressure to obtain the crude substance. Obtained 550mg.
This crude material was again placed on a silica gel column (Wako Gel C-200, manufactured by Wako Pure Chemical Industries, Ltd., 40 g) and developed with a developing solvent benzene:ethyl acetate (88:12, v/v). In the same manner as above, a portion of the eluate was subjected to thin layer chromatography, and fractions containing a substance exhibiting an Rf value of 0.65 were collected and concentrated to dryness under reduced pressure to obtain 107 mg of yellow powder. When this was analyzed, it was confirmed to be bethenone C.

[Brief explanation of drawings]

The drawing shows the compound of the present invention from the starfish gastrulae.
FIG. 2 is a diagram showing inhibition of DNA, RNA, and protein synthesis.

Claims (1)

[Claims] 1 formula A novel compound bethenone C represented by 2 Belongs to poma betae, formula 1. A method for producing the novel compound Bethenone C, which comprises culturing a microorganism capable of producing the novel compound Bethenone C shown in the formula in a medium to produce and accumulate the compound, and then collecting the same.