JPH0466582A - Macrolide substance - Google Patents

Abstract

NEW MATERIAL:A compound shown by the formula having the following physical and chemical properties. Molecular weight: 594. Molecular formula: C22H50O10. Specific rotatory power: [alpha]D=+84.4 deg. (C=0.26, CHCl3). Mass spectrometric analysis (FABMS): m/z 700 [(M+DEA+H)<+>]. Solubility: soluble in chloroform, ethanol and methanol and insoluble in water. USE:An antitumor agent. PREPARATION:Mycal sp. is extracted with ethanol, the extracted solution component is distributed into two layers of water and diethyl ether and the objective substance is isolated from the transferred component of the diethyl ether layer.

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial Application Field) The present invention relates to a novel macrolide substance that is an extracted component from a sponge and exhibits antitumor activity.

(Prior Art) Several physiologically active substances have been isolated from sponges. For example, Ha
richin (Richondria okadai), a 38-carbon fatty acid (okadaic acid, 0k
adaic acid) and norhalichondrin [Journal of the American Chemical Society (J, Am, Chem, Soc, )]
+ 1981103 2469-2471 and J, Hachim, Chem, Soc,, 1985107 479
6-47981. Pandaros acanthifolium (Pa
ndaros acanthifolium), polyether carboxylic acid with episulfide bonds (acantifolicin, Acanthifolicin)
reported jJ, Am.

Chem, Soc, 1981.103.2467-
2468], and from the Caribbean sponge Tedani aignis (Tedania 7), the cytotoxic 18
Ring-membered macrolide (tedanolide)
e) reported [J, Am, Chem.

Soc, 19B4.106.7251-7252].

(Structure of the Invention) The present inventors searched for physiologically active substances from sponges produced in Japan, and discovered the Subirokecool compound, which exhibits antitumor activity from the sponge Discodermia caJ, and have previously filed a patent application for the compound. Publication No. 62-178595). After further exploration, I found Kuchifuri Island (Shikoku)
My-, a sponge that lived at a depth of 10 to 20 meters in the surrounding sea area.
A substance exhibiting strong cytotoxicity was found in the lipophilic extract components of 9a order sp, and this substance was isolated to complete the present invention.

That is, the present invention relates to a macrolide substance represented by chemical structure 2 [1].

(Manufacturing method) Macrolide substances are obtained by extracting cotton wool "ya]±sp" with ethanol, and diluting the extracted components with water and diethyl ether.
It is possible to separate the layers and select from among the components that migrate to the diethyl ether layer.

The raw material, Nigori cotton jaw 1 es p, grows in the seas around Shikoku and near Japan, and can be collected and used. The whole cotton is crushed and then extracted.

Extraction is performed at room temperature, and the ethanol used for extraction is 99% or higher. Extraction is performed several times using ethanol at least twice the amount of the sponge. After concentrating the extract under reduced pressure if necessary, it was subjected to two-layer partitioning with water-diethyl ether, the diethyl ether layer was partitioned into two layers with hexane and 90% methanol, and the 90% methanol-soluble components were separated by flash chromatography (silica gel). to separate the active ingredient. The active ingredient thus obtained is further purified using reverse phase high performance liquid chromatography to obtain the pure macrolide substance.

The physicochemical properties of the purified macrolide substance are as follows.

]) Molecular weight 594 2) Molecular formula C1□11. ．． 0. .

3) Optical rotation iαlo +84.4° (C=0.26.
C11 ([3)4) Infrared absorption spectrum (IR)
(film)3450, 3000.2950.175
5.1710.1455°1370, 1240.121
0.1140.1115.1090°1035, 980
．． 890.860.830.810.760゜670
cm' 5) Mass spectrometry (FABMS) TiL700 [(M+
DEA+H)"] 6) 'HIIMR, mouth CNMR (
5001'1) lz) Table 1 7)? 8 Solubility Chloroform: Soluble Ethanol: Soluble Methanol: Soluble Water: Insoluble Table 1 arbon I″Cl 73.5s 72.5d 85.1d 49.3d 217.3s CD, OD H 3,686 (1, 9) 3.60dd (+, 9° 3.12m (7,1 C 171,6s 71, Od 9.6) 83.3d 9.6) 48.2d 2+5.1s CDCI!, 3 H 3,68bd ( 7,5) 3.60dd (1,5,9,5) 3.05m (7,1,9,5) arbon 51.1d Bo, 2d 138.4s 130.1d 46.1d 213.8s 3B, 6t 26, Ot 49.56 15 215.6s 53.2d 77.7d D30D 3.11m (7,0,10,1) 3.9sd (10,1) 5.26dd (1,3,9,7) 3 .41m (6,99°7) 2.30dd (7,8,7,8) 1.57m (3,17,8 13,8) 1.95m (7,8,9,0°13.8) 2.77m (3,1,7,1°9.0) 3.35m (4,3,10,3 11,9) (10,3) 3.17d CDCff.

49.5d 3.05m (6,9,10,0)7
9.4d 4.07d (10,0) 136.3s 129.9d 5.39dd (1, 1, 8, 3) 45
, Od 3.27m (6,9,8,3)210.
8s 37.5t 2.22m 24.6t 1.98m 1.64m 4B, 1d 2.71m 213.6s 50.8d 3.27m 76.6d 3.23d carbon 64.0s 67.4d 32.4d 131.8d 126.0d 13.5q 15.2q 15.6q 10.5q 16.2q 15.7q 65.7t 11.4q 16.7q 61.1q Ih0D 2.656 (9,3) 2.47m (7,9, 10,5) 5.32ddd (1,7°10.5. 10.9) 5.46m (6,8,10,9) 1.61dd (1,7,6,8) 1.22d (7, 1) 1.26d (7,0) 1.636 (1,3) 1,OOd (6,9) 1.116 (7,1) 4.01dd(10,5,11,9) 4.17dd( 4,3,10,5) 1.43s 1.096(7,9) 3.33s 62.7s 66.76 31.1d 130.1d 125.0d 13.3q 14.5q 15, 1q 10.0q 16 .1q 15.2q 64.5t 11.2q 1B, 4q 60.5q CDCf.

2,60d (9,3) 2.43d (6,9,10,5) 5.23ddd (1,6,10,5 10,5) 5.45m (6,8,10,5) 1. 57dd (1,6,6,8) 1.216 (7,1) 1.28d (6,9) 1.60d (1,1) 1.03d (6,9) 1.11d (7,3) 4.136d (5,3,10,7) 1.33s 1.10d (6,9) 3.29s (Utility) The compound of the present invention has a poor antitumor effect. The in vitro tumor cell proliferation inhibitory effect and the in vivo antitumor effect will be shown below along with experimental methods.

(i) In vitro effect experiment method Tumor cells shown in Table 2 were treated with RP containing 10% newborn serum.
Using the solution added to the MI 1640 culture solution, adjust the number of cells in the culture solution to 1 x 10 cells/family. Dispense one goose into a plastic well. The macrolide substance is dissolved in dimethyl sulfoxide (hereinafter abbreviated as DMSO), and added to the cell suspension culture solution so that the final concentration of DMSO is 0.4% by volume and the macrolide substance is at the specified concentration, and then 5% carbonate is added. The cells were cultured in gas-containing air for 3 days. As a control, a cell suspension culture solution to which 4% by volume of DMSOO was added was similarly cultured.

After culturing, the cells were stained with Trihan Blue staining solution, the number of viable cells was counted, and the IC5° value (50% cell growth inhibitory concentration) was determined from the inhibition rate relative to the control. Mitomycin C was used as a control drug. The results are shown in Table 2.

Table 2 Ta. The results are shown in Table 3.

T/C%-T/CX100 T: Median survival days of administration group C: Median survival number of control group P38B < 0.000094
0.011L1210 0.00021
0.057 to 562 0.
00018 0.020Heta S
3 0.00033 0.02
1KB 0.00022
< 0.0156(ii) in
Antitumor test against leukemia cell P388 in vivo DBA/CRJ mice 4 x 105 7-day-old P388 cells were intraperitoneally transplanted into a group of 5 BDF,
/CRJ mice (5 weeks old, male) were transplanted intraperitoneally. The active ingredient was intraperitoneally administered in a predetermined amount according to the schedule shown in Table 3 starting 24 hours after transplantation. The median survival days of each active ingredient administration group was determined for the tumor growth suppressive effect, and the survival rate (T/C%) of each administration group relative to the control group was calculated using the following formula. For use, the macrolide substance is administered in a form convenient for producing an anti-tumor effect.3 Although the macrolide substance may be a medicament in its raw form, it is mixed with pharmaceutically acceptable diluents and/or in accordance with pharmaceutical practice. Alternatively, it may be provided in a composition as a mixture with other pharmacologically active substances. Therefore, the macrolide substance of the present invention can be appropriately administered in a form for oral or parenteral administration. For example, powders, granules, tablets, sugar-coated tablets,
Capsule, bill, planing, suspension, liquid, emulsion, injection,
It is an aerosol agent.

The dosage of the macrolide substance of the present invention varies depending on various factors such as sensitivity differences, age, sex, body weight, administration method, administration timing, interval, medical condition, physical condition, properties of the pharmaceutical preparation, type of adjustment, etc. do.

(Example) Next, the compound of the present invention and the method for producing the same will be further explained with reference to Examples.

Example 1 1.9 kg of a sample of a sponge (machi ca, , , , je sp,) collected from the sea area around Kuchifuri Island, Shikoku (water depth 10 to 20 m), which had been cryopreserved at -20°C, was treated with ethanol ( 61)
The mixture was extracted three times and partitioned into two layers with water and diethyl ether. Furthermore, the diethyl ether layer was partitioned into two layers with hexane and 90% methanol, and the 90% methanol layer was fractionated by flash chromatography (Kieselgel 60H). The active fraction eluted with chloroform/methanol was gel-filtered (TOYOPEARl, HW40SF
, 1 coform/methanol (1:1) and further purified by reverse phase flash chromatography (YMC-O
IIS). Finally, the active fraction eluted with 80% methanol was analyzed by HPLC (CAPCELL PAK C,
, 20 mmφX 250 mm) to obtain compound 105.2 of the present invention.

Claims (1)

[Claims] formula ▲Contains mathematical formulas, chemical formulas, tables, etc.▼ A macrolide substance indicated by .