JP3074084B2 - Novel 16-membered macrolide derivatives and production method - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention
The present invention relates to a 6-membered ring macrolide derivative, a method for producing the same and a use thereof.

[0002]

2. Description of the Related Art Macrolide antibiotics effective against gram-positive bacteria, mycoplasma, chlamydia, etc. are classified as clinically important antibacterial agents because they can be administered orally and have low toxicity. Myokamycin (MOM), which has better pharmacokinetics and less bitterness than natural compounds
(Journal of Antibiotics, 29 (5), 536 (1
976), Japanese Journal of Antibiotics, 35 (6), 1462 (1982)) has been actively used in clinical practice as a semi-synthetic 16-membered macrolide antibiotic. The present inventors have selected 16-membered macrolide antibiotics as research subjects because of their low resistance induction, little interaction with other drugs, and little effect on the intestinal tract. We have been conducting synthetic and biochemical exploration research on compounds effective against bacteria.

[0003] From these studies, a biochemical method for specifically cleaving the acyl group bonded to the 3-position hydroxyl group of a 16-membered macrolide with a certain mold has been invented as a means for enhancing the in vitro antibacterial activity. and Patent application (JP-a
No. 6-16691) . Next, as a technique for improving the pharmacokinetics in mouse animal experiments, a technique for converting an oxo group bonded to the 9-position of a 16-membered macrolide into a hydroxyl group using a certain actinomycete was disclosed (JP-A-6-206897). ) . Also,
As a result of repeated synthetic chemistry studies aimed at 16-membered macrolide derivatives that are stable in vivo and are unlikely to have reduced antibacterial activity even when metabolized, the two hydroxyl groups of the mycalose moiety of the 16-membered macrolide derivative are both We succeeded in newly synthesizing a derivative linked by an ether with two alkyl groups, and filed a patent application (JP-A-6-9679) .

[0004] By the way, 14-membered ring and 16
An overview of membered macrolide antibiotics reveals interesting structural facts. That is, in the case of a 14-membered macrolide, 3
As a neutral sugar that binds to the hydroxyl group at the position, cladinose is mainly used, and L-sugars having various structures such as mycalose and oleandose are known. On the other hand, in the case of a 16-membered ring macrolide, the neutral sugar that binds to the hydroxyl group at the 4-position of micaminose, that is, the 4′-position, is mainly L-mycarose and its 4-O-acyl form, and L- cladinose is known. However, there is no known natural product having an α-glycosidic bond to the 4′-hydroxyl group, and therefore, I was interested in the synthesis and antibacterial activity of the compound. To elucidate some of these interests, a carbomycin B derivative (16-membered macrolide) in which 4-O-isovaleryl-L-clazinose is α-glycosidically linked to the 4′-hydroxyl group has been synthesized (Chemistry Letters). , 769 (1977)). It is reported that its antibacterial activity was enhanced against acid-fast bacterium as compared to Carbomycin B. On the other hand,
Derivatives from which the isovaleryl group attached to the 4 "hydroxyl group has been removed are not yet known. By comparison, the antibacterial activities of erythromycins A and C and B and D against Bacillus subtilis show that A is greater than C than D. B is also known to have strong antibacterial activity (Journal of the
American Chemical Society, 99 (5), 1620 (197
7)). That is, in erythromycins, it is suggested that the methyl group bonded to the hydroxyl group at the 3-position of cladinose plays an important role in enhancing the antibacterial activity.

[0005]

It has been reported that a 16-membered macrolide antibacterial agent having few side effects generally has a weaker in vitro antibacterial activity than that of a 14-membered ring (Antimicrovial Agents & Co., Ltd.).・ Chemotherapy, 32 (11), 1710 (1988)). Therefore, Streptococ , one of the main causative bacteria of upper respiratory tract infections that are important in clinical
The emergence of 16-membered macrolide antibiotics with enhanced antibacterial activity against the genus cus is expected.

[0006]

Means for Solving the Problems The inventors of the present invention have conducted synthetic and biochemical studies in order to meet the above-mentioned expectations, and have first applied a patent application (Japanese Patent Laid-Open No. 6-9679) to glycosyl. A 16-membered macrolide derivative in which 4-O-propionyl-L-cladinose was α-glycosidically bonded to the hydroxyl group at the 4′-position was produced by using a synthetic technique for a 16-membered macrolide derivative via the conversion. By treating this with a certain microorganism or the like or an enzyme produced by a living body, for example, by microbial conversion using a strain of fungus PF1108 strain, it was bonded to the hydroxyl group at the 4 "position. The propionyl group was cleaved efficiently, and unmodified L-clazinose was linked to α-glycosidic bond at the 4-position, ie, the 4′-position, of the thus obtained mycaminose.
The present inventors have found that a 6-membered macrolide derivative strongly inhibits the growth of clinically important Gram-positive bacteria, particularly Streptococcus sp., Thereby completing the present invention.

The gist of the present invention is to provide a compound represented by the following formula (I):

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Wherein R ¹ is an oxygen atom , R ² is a hydrogen atom or a group of the formula COR ⁴ (where R ⁴ is a linear alkyl group having 1 to 3 carbon atoms), and R ³ is a hydrogen atom or A group of the formula COR ⁴ (where R ⁴ has the same meaning as described above) is shown. Or a pharmaceutically acceptable salt thereof. The compound represented by the general formula (I) according to the present invention is produced as follows by the method shown in the flow chart. In this case, the cladinoside
Although the hydroxyl group at the 4-position is protected by a propionyl group, it is also within the scope of the present invention to carry out this step with protection using another acyl group such as a butyryl group. In addition, the first position of cladinose uses an ethyl glycosyl form, but other glycosyl forms,
For example, performing the step by using a methylglycosyl compound or the like is also included in the scope of the present invention.
Although a compound in which the hydroxyl group at position 3 of the macrolactone ring is propionylated is used, the step may be performed using a compound acylated with another acyl group, for example, an acetyl group or a butyryl group. It is also included within the scope of the present invention. That is, via the compound (7) shown in the process diagram 1 and analogous substances thereof, for example, a derivative having a different acyl side chain bonded to the hydroxyl group at the 3-position and / or 4 "-position present in the structural formula, etc. A method for producing the compound represented by the general formula (I) using a chemical method or a synthetic chemical method is also included in the scope of the present invention.

[0009]

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First, a method for synthesizing a sugar component donor in a glycosylation reaction will be described. First, an appropriate acyl side chain is introduced into a free hydroxyl group of L-clazinoside, for example, compound (1) (ethyl β-L-clazinoside) to give 4-O-acyl.
-Synthesize L-clazinoside. The acyl side chain may be a lower linear or branched acyl group, for example, an acetyl group, a propionyl group, a butyryl group, a valeryl group, an isovaleryl group and the like. That is, in the presence of an organic or inorganic base, L-
The 4-position acyl compound is obtained by reacting cladinoside with an acylating reagent such as a corresponding acid anhydride or acid halide. For example, compound (1) is reacted with an equimolar or excess amount of propionic anhydride in the presence of 4-dimethylaminopyridine to give compound (2). In addition, although all of the ethyl glycosides in the present process diagram 1 are shown as β-forms, no inconvenience will occur in the subsequent reaction even if α-glycoside is used alone or mixed with β-form.

Next, 4-O-acyl-L-cladinoside, for example, compound (2) is acid-hydrolyzed in a water-containing solvent to quantitatively obtain 4-O-acyl-L-cladinose, for example, compound (3). Preferred acid hydrolysis conditions are a mixture of 0.5-1N hydrochloric acid and acetonitrile or 1,4-dioxane, and the reaction proceeds smoothly at 0 ° C to 40 ° C, preferably at 20 ° C to 30 ° C. is there. It should be noted that there is another report on the synthesis method of 4-O-acyl-L-clazinose (Chemistry Letters, 769 (1977)).

The obtained 4-O-acyl-L-clazinose, eg, compound (3), is converted to a sugar component donor, eg, compound (4)
Convert to In recent years, glycosylation methods have been actively developed. In addition to conventional glycosylation methods using 1-halogen sugars (bromo sugars, chloro sugars), 1-thio sugars, 1-fluoro sugars, Recently, attention has been paid to a method for introducing a sugar chain using an acyl sugar as a sugar component donor. 4-O-acyl-L-cladinose, for example, compound (3) can be converted to the useful sugar component donors described above such as 1-thiosugar, 1-fluorosugar, 1-acylsugar and the like. Yes, and at the same time glycosylation using them is applicable to the preparation of the compounds of the general formula (I) according to the invention.

As an example for producing a useful sugar component donor, 1-thio-branched sugar (Journal of Chemical Co., Ltd.)
Society Parkin Transaction I, 1989 ,
799) will be described. 4-O-acyl-L-clazinose, for example, compound (3) is converted to aldrithiol-2 (2,2 '
-Dipyridyldisulfide) and tri (n-butyl) phosphine to give 1- (2-pyridylthio) sugars, for example compound (4). The thiosugar is obtained as a mixture of α-thioglycoside and β-thioglycoside in approximately equal amounts. However, at present, the effect of the configuration at the 1-position of the 1-thiosugar on the configuration of the anomeric carbon of the glycosylation product has not always been completely elucidated. Therefore, the two anomers of the 1-thiosugar may or may not be separated and may be used as a mixture in the glycosylation reaction. The 1-thiosugar may be a phenylthiosugar, a methylthiosugar, or a thiosugar bonded to a heterocyclic ring other than a pyridine ring, in addition to the aforementioned 1- (2-pyridylthio) sugar.

Second, the introduction of 4-O-acyl-L-clazinose and the reduction of dimethylamino group N-oxide will be described. 3-O-acyl-9-dehydro-16-membered macrolide N-oxide derivatives deficient in neutral sugars, for example, a known substance, midecamycin A ₃ (Journal of Antibiotics, 24 (7), 476 ( 1971)) to the compound (5) (JP-A-6-9679) , which is produced through three steps, is combined with the above-mentioned 1-thiosugar of 4-O-acyl-L- clazinose in the presence of silver perchlorate, for example, Reacting compound (4),
The desired α-glycoside, for example, compound (6) is obtained. As an activator for the condensation reaction, in addition to silver perchlorate, silver triflate, lithium perchlorate, trityl perchlorate, iodine-trityl perchlorate, silver carbonate, N-iodosuccinimide, methyl triflate Rate, phenylselenyl triflate, dimethyl (methylthio) sulfonium triflate and the like can be used alone or in combination. The solvent may be acetonitrile, propionitrile, methylene chloride, benzene, toluene, dioxane, tetrahydrofuran, or the like, and is preferably acetonitrile. The reaction proceeds at -20 ° C to 30 ° C.
In this glycosylation reaction, a small amount of β-glycoside is by-produced depending on the reaction conditions.However, since the Rf value by silica gel thin-layer chromatography is clearly different from that of α-glycoside, it can be easily separated by the chromatographic method. Is possible.

With regard to the glycosylation reaction using 4-O-acyl-L-clazinose as a sugar component donor, for example, compound (4), the sugar component acceptor is limited to compound (5) only. It is also possible to use 9-dehydro-demycarosylplatenomycin (DDM-PLM) (Journal of Antibiotics, 28 (10), 789 (1975)) without any need. However, as a sugar component donor, 1-
When (2-pyridylthio) sugar is used, the glycosylation yield is more excellent when compound (5) is used.
By the way, the compound (5) has a protected dimethylamino group and does not require deprotection of the aldehyde group at the 18-position after the desired chemical transformation. Is a very useful synthetic intermediate. Further, in order to produce the compound represented by the general formula (I) in the present invention, as the sugar component acceptor, the hydroxyl group at the 3-position of the compound (5) is not a propionyl group but an acetyl group or a butyryl group. Analogues of the compound (5) combined with the compound (5) may also be used. On the other hand, a 16-membered macrolide derivative
When introducing 4-O-acyl-L-clazinose to the 4'-hydroxyl group, a synthesis example using a strategy using a glycal derivative as a sugar component donor (Carbohydrate Research, 54 , 85 (1977)) It has been reported (Chemistry Letters, 769 (1977)).

Next, α-glycoside (coupling product),
For example, the compound (6) is reacted with phosphine or phosphite to obtain a derivative obtained by converting a dimethylamino group N-oxide to a dimethylamino group, for example, a compound (7). N-
The reducing agent for the oxide may be a phosphine such as triphenylphosphine or tri (n-butyl) phosphine, or a phosphite such as trimethylphosphite or triethylphosphite. As a solvent, chloroform, methylene chloride, ethyl acetate, ether, benzene and the like may be used. Compound (7) itself is a novel substance and has excellent antibacterial activity against Gram-positive bacteria.

Third, 4'-O- having a dimethylamino group
The (4-O-acyl-α-L-clazinosyl) derivative, for example, the acyl group of the cladinose moiety of the compound (7) is cleaved by a known biochemical method to obtain a compound represented by the general formula (I) of the present invention.
R ¹ is represented by an oxygen atom, R ² is represented by a group of the formula COR ⁴ (where R ⁴ is a straight-chain alkyl group having 1 to 3 carbon atoms), and R ³ is a 16-membered ring represented by a hydrogen atom. A macrolide, for example compound (8), is obtained. The technique of cleaving an acyl group bonded to a 4 "-hydroxyl group in a 16-membered ring macrolide compound by a biochemical method is known per se, and there are many reported examples (Chemistry and Biology, 8
(3), 139 (1970), and Fermentation and Industry, 37 (8), 749 (1979)). However, there has not yet been reported an example in which the microbial conversion of a 16-membered macrolide compound in which the hydroxyl group at the 3 "-position is methylated has been carried out. The microorganism conversion by the PF1108 strain, that is, 4 ″ in the 16-membered macrolide compound
The conversion which cleaves the acyl group bonded to the hydroxyl group at the position is achieved. On the other hand, microorganisms having such functions and characteristics are PF
The present invention is not limited to strain 1108, and it is possible to utilize enzymes produced by other microorganisms or living organisms, such as certain molds or actinomycetes (Japanese Patent Application Laid-Open No. 48-72389). The microbial conversion method using the PF1108 strain is characterized in that it does not require complicated control of an enzyme system or strict adjustment of the hydrogen ion concentration. The details of the present microorganism conversion method are described in Examples described later.

The PF1108 strain used in the microbial conversion was 1990
It is a fungus newly isolated from the leaves of a plant (colored leaves) in Kyoto City, Kyoto Prefecture in April. The bacteriological properties are as follows. (1) Characteristics on each medium It grows well on Czapek yeast extract agar medium, and becomes a velvety, flat colony with a diameter of about 35 mm when cultured at 25 ° C for 7 days. Its color tone is yellow-orange and ocher, and the back is ocher. It grows well on malt extract agar medium, and becomes a slightly rope-like colony with a diameter of about 30 mm when cultured at 25 ° C for 7 days. Its color tone is white to cream, and the back surface is dull yellow. It grows well on cornmeal agar medium, and becomes a velvety to wool-like, flat colony with a diameter of about 35 mm when cultured at 25 ° C for 7 days. Its color is grayish green and the back is gray. On a Tzapek-Docs agar medium, it hardly grew after culturing at 25 ° C. for 7 days. The soluble pigment and secretion did not grow on any medium. The culture at 37 ° C. did not grow on any medium.

(2) Morphological characteristics Observation under a microscope showed that no morphology showing mycological characteristics, such as conidia, was observed on any medium, with only hyphae. In addition to the above media, potato dextrose agar,
After culturing for 1 month in each medium such as an oatmeal medium and a potato carrot medium, no conidia were observed. Based on the above bacteriological properties, it is considered that the present strain PF1108 belongs to the azoosporidious incomplete fungi. This strain has been deposited with the Institute of Biotechnology, Institute of Biotechnology, Japan as a non-spore-free incomplete bacterium PF1108 strain (deposit number FERM P-13363).

Next, the neutral sugar moiety is an α-L-clazinosyl residue, and a 16-membered macrolide derivative having an acyl group at the hydroxyl group at the 4-position, ie, the 4 ″ -position, is used as an important intermediate. The production method for obtaining the deacylated product will be described in detail below. To carry out the present microorganism conversion method, first, the microorganism is pre-cultured in a liquid medium. A 16-membered macrolide compound may be added to a separately prepared liquid medium, and then the preculture may be inoculated. Inoculation of the preculture may be performed at any time after the growth of the microorganism.
It is preferable to inoculate around 4 hours. The amount of the substrate to be added is usually 0.01 mg / ml to 2 mg / ml.

As a nutrient source of the medium, known ones conventionally used for mold cultivation can be used. Preferably,
It is preferable to use glucose and starch as the carbon source, and soybean meal and wheat germ as the nitrogen source. In addition, if necessary, it is possible to add sodium, potassium, calcium, magnesium, cobalt, chlorine, phosphoric acid, sulfuric acid, and other inorganic salts capable of generating ions. In addition, organic substances and inorganic substances that promote the growth of bacteria and promote the conversion of the substrate can be appropriately added.

As the culture method, a culture method under aerobic conditions,
In particular, the submerged culture method is most suitable. The appropriate temperature for culture
Culture at 22-30 ° C, but often around 26 ° C. The conversion efficiency of the substrate varies depending on the medium and the culturing conditions, but the maximum accumulation of the converted substance usually occurs in 24 to 240 hours in both shaking culture and tank culture. When the accumulated amount of the conversion substance in the culture solution reaches the maximum, the culture is stopped, and the target substance is isolated and purified from the culture solution.

In order to obtain the desired conversion substance from the above-mentioned culture solution, the cells are removed after the culture, the culture solution having no solid content is made alkaline, and then an organic solvent which does not mix with water, such as butanol and ethyl acetate. By performing extraction with chloroform, methylene chloride, or the like, the converted substance is retained in the organic solvent layer. To further purify the converted substance, chromatography using an adsorbent such as silica gel or alumina may be performed. For small-scale purification, use preparative TL
It is effective to use C. By the way, the method for purifying the conversion substance is not limited to the method using the above-mentioned adsorbent, but includes gel filtration, countercurrent distribution chromatography,
It is possible to apply a method which can be generally used when purifying a natural or synthetic organic compound. Further, the conversion substance obtained in the form of a free base may be converted into the corresponding acid salts by a conventional method using a pharmaceutically acceptable inorganic or organic acid. Accordingly, acid salts thereof as well as free base converters are intended to be included within the scope of the present invention.

[0024]

The compound (8) represented by the general formula (I) obtained by the present invention has a strong antibacterial activity against clinically important gram-positive bacteria, particularly Streptococcus , as shown in Table 1. doing. For comparison, DOP in which the α-L-clazinosyl moiety of compound (8) was replaced with an α-L-mycarosyl residue
The antibacterial activity (simultaneous measurement) of the substance (JP-A-48-10288) (9-dehydro-3-O-propionylleucomycin V) was shown.

Table 1: Antibacterial activity of the compound of the present invention (MIC; μg / ml) Test bacterium Compound of the present invention Comparison (8) DOP substance S. aureus 209P JC-1 0.20 1.56 S. aureus M133 0.78 3.13 S. aureus M126 >100> 100 S. aureus MS15009 / pMS99>100> 100 S. aureus MS15026>100> 100 S. aureus MS15009 / pMS98 0.39 3.13 S. aureus MS15027 0.78 3.13 S. epidermidis ATCC14990 0.78 3.13 M. luteus ATCC9341 <0.025 0.20 E faecalis W-73 0.39 3.13 E. coli NIHJ JC-2>100> 100 K. pneumoniae PCI602>100> 100 S. pneumoniae IP692 0.05 0.39 S. pneumoniae Type I <0.025 0.20 S. pyogenes Cook 0.10 0.39 B. catarrhalis W -0500 1.56 12.5 B. catarrhalis W-0506 0.78 12.5 H. influenzae 9334 1.56 50 H. influenzae Type b 12.5 100

Examples for obtaining the compound of the present invention and physicochemical properties of the compound of the present invention will be described below. According to the present example, the neutral sugar moiety is an α-L-cladinosyl residue, and the neutral sugar moiety has an acyl group at the 4-position, that is, the hydroxyl group at the 4 ″ -position.
The usefulness of using a 6-membered macrolide derivative as an important intermediate to obtain the corresponding 4 "-deacylated derivative using certain molds was demonstrated. It is possible to devise various methods for producing the substance by using the same kind of biochemical method by using it as a simple intermediate, while it is also possible to devise various methods for the deacylation reaction by using a synthetic chemistry method. The present invention is not limited to the examples, and the known means can be modified based on the properties of the compound (8) represented by the general formula (I) revealed by the present invention, as well as the means for modifying the examples. It encompasses all methods of synthesis, production, extraction and purification, applied alone or in combination.

[0027]

EXAMPLES Example 1 Compound (2) (ethyl 4-O-propionyl-β-L-
Production method of cladinoside) Compound (1) (JP-A-6-9679)
To 6.69 g, 200 ml of anhydrous pyridine was added and dissolved, and 10.2 g of propionic anhydride and 2.00 g of 4-dimethylaminopyridine were added, followed by heating at 50 ° C. for 80 minutes. After returning the reaction solution to room temperature,
10.0 g of water was added and left at the same temperature for 30 minutes. Next, the reaction solution was concentrated under reduced pressure, and the obtained residue was extracted with 600 ml of chloroform. The chloroform layer was washed twice with a 5% aqueous potassium hydrogen sulfate solution (600 ml each) and a saturated aqueous sodium hydrogen carbonate solution (600 ml).
, And once with 600 ml of saturated saline, and then the organic layer was dried over anhydrous sodium sulfate and filtered. The residue obtained by concentrating the filtrate under reduced pressure was subjected to silica gel column chromatography (700 g, toluene-ethyl acetate (10: 1
1) → Purification (5: 1)) to obtain 6.50 g of compound (2). Physicochemical properties of compound (2) (1) Color and shape: colorless oil (2) Molecular formula: C ₁₃ H ₂₄ O ₅ (3) Mass spectrum (SIMS): m / z 261 (M + H) ⁺ (4) Specific rotation: [α] _D ²⁵ -1 ° (c1.0, CHCl ₃ ) (5) ¹ H NMR spectrum (400 MHz, CDCl ₃ ) δ (pp
m): 4.71 (dd, 1-H), 1.51 (dd, 2-Hax), 2.20 (dd, 2-
Heq), 4.68 (d, 4 -H), 3.93 (dq, 5-H), 1.15 (d, 6-H 3),
1.13 (s, 7-H ₃ ), 3.52 (dq, 1-OC H ₂ CH ₃ ), 3.97 (dq, 1-OC H
_{2 CH 3), 1.23 (t} , 1-OCH 2 C H 3), 3.29 (s, 3-OCH 3), 2.41 (q,
4-OCOC H ₂ CH ₃ ), 2.42 (q, 4-OCOC H ₂ CH ₃ ), 1.17 (t, 4-OCOC
H ₂ C H ₃ )

Example 2 Compound (4) (1-deoxy-4-O-propionyl-1- (2-pi
Production Method of Lysylthio) -L-clazinoside) To 6.00 g of compound (2), 120 ml of 1,4-dioxane was added and dissolved. 120 ml of 1N hydrochloric acid was added thereto, and the mixture was stirred at room temperature for 15 minutes, and then allowed to stand at the same temperature for 2 days. 240 ml of a saturated aqueous sodium hydrogen carbonate solution was gradually added to the reaction solution, and the mixture was extracted with 200 ml of methylene chloride. Add methylene chloride to the aqueous layer again.
0 ml was added for extraction. The methylene chloride layers were combined and washed twice with 100 ml each of saturated saline. The methylene chloride layer was dried over anhydrous sodium sulfate and filtered, and the filtrate was concentrated under reduced pressure to obtain 4.95 g of compound (3) (4-O-propionyl-L-clazinose). 4.95 g of compound (3) was dissolved in 90 ml of anhydrous methylene chloride and cooled to 0 ° C. On the other hand, 7.51 g of aldrithiol-2 was dissolved in 90 ml of the same solvent, 11.5 ml of tri (n-butyl) phosphine was added, the mixture was cooled to the same temperature, and then added dropwise to the above solution. After reacting at room temperature for 4 hours, the reaction solution was concentrated under reduced pressure, and the obtained residue was purified by silica gel column chromatography (1.0 kg, chloroform-ethyl acetate (100: 1)) to obtain 5.84 g of compound (4). Obtained. Further, the α- and β-thioglycosides of compound (4) are separated by using preparative TLC (developing system: n-hexane-chloroform-ethyl acetate (5: 6: 1), developed twice). It was possible to do. Compound (4) Physicochemical properties of α-thioglycoside (1) Color and shape: colorless needle crystals (2) Molecular formula: C ₁₆ H ₂₃ NO ₄ S (3) Mass spectrum (FDMS): m / z 325 (M ) ⁺ (4) Specific rotation: [α] _D ¹⁸ -330 ° (c1.0, CH ₃ O
H) (5) Melting point: 112-113 ° C. (6) ¹ H NMR spectrum (400 MHz, CDCl ₃ ) δ
(ppm): 6.39 (br d, 1-H), 2.19 (dd, 2-Hax), 2.46 (b
rd, 2-Heq), 4.75 (d, 4-H), 4.48 (dq, 5-H), 1.11 (d,
6-H ₃ ), 1.18 (s, 7-H ₃ ), 3.34 (s, 3-OCH ₃ ), 2.44 (q, 4-O
COC H ₂ CH ₃ ), 2.45 (q, 4-OCOC H ₂ CH ₃ ), 1.19 (t, 4-OCOCH ₂ C
H ₃ ), 7.29 (dt, 3'-H), 7.51 (dt, 4'-H), 7.01 (ddd, 5'-H
H), 8.47 (ddd, 6'-H) Compound (4) Physical and chemical properties of β-thioglycoside (1) Color and shape: colorless needle-like crystals (2) Molecular formula: C ₁₆ H ₂₃ NO ₄ S (3) Mass spectrum (FDMS): m / z 325 (M) ⁺ (4) Specific rotation: [α] _D ¹⁸ + 56 ° (c1.0, CH ₃ OH) (5) Melting point: 101-102 ° C (6) ¹ H NMR spectrum (400 MHz, CDCl ₃ ) δ
(ppm): 5.69 (dd, 1-H), 1.85 (dd, 2-Hax), 2.42 (d
d, 2-Heq), 4.74 (d, 4-H), 4.13 (dq, 5-H), 1.15 (d, 6-
H ₃ ), 1.18 (s, 7-H ₃ ), 3.33 (s, 3-OCH ₃ ), 2.43 (q, 4-OCO
C H ₂ CH ₃ ), 2.44 (q, 4-OCOC H ₂ CH ₃ ), 1.19 (t, 4-OCOCH ₂ C
H ₃ ), 7.31 (br d, 3'-H), 7.55 (dt, 4'-H), 7.05 (br dd,
5'-H), 8.45 (br dd, 6'-H)

Example 3 Preparation of compound (7) (3 "-O-methylmidecamycin A ₃ )
Method 3.47 g of highly dried compound (4) and compound (5) 1.
To 11 g, 32 ml of anhydrous acetonitrile was added and dissolved. Activated powdery molecular sieves 4A (12.0 g) was added thereto, followed by stirring at room temperature for 15 minutes. After cooling the reaction mixture to −15 ° C., 3.68 g of anhydrous silver perchlorate was added, and the mixture was stirred at the same temperature under light shielding conditions. The reaction temperature was gradually returned to room temperature over 2 hours, and then the mixture was stirred at room temperature for 24 hours under light-shielded conditions. Separately 1.0 l of methylene chloride and saturated aqueous sodium hydrogen carbonate solution 1.0
While the l were combined and vigorously stirred, the previous reaction mixture was instantaneously charged and vigorously stirred for 30 minutes. After filtering the insoluble matter, the methylene chloride layer was separated and the aqueous layer was methylene chloride 500 m each.
Re-extracted twice with l. Combine the organic layers and add saturated saline 1.2 l
After washing with, a methylene chloride layer was separated, dried over anhydrous sodium sulfate, and filtered. The residue obtained by concentrating the filtrate under reduced pressure was subjected to silica gel column chromatography (150
g, chloroform-methanol (50: 1))
102 mg of crude compound (6) (3 "-O-methylmidecamycin A ₃ N-oxide) was obtained. This was dissolved in 9.0 ml of anhydrous methylene chloride without further purification, and 700 mg of triphenylphosphine was added. The reaction was performed at room temperature for 4 days, and the residue obtained by concentrating the reaction solution under reduced pressure was subjected to preparative TLC (developing system:
Purification with chloroform-methanol (50: 1, developed twice) gave 48 mg of compound (7). Physicochemical properties (1) Color and shape of the compound (7): colorless solid (2) Molecular _{formula: C 42 H 67 NO 15 (} 3) Mass spectrum (FDMS): m / z 826 (M + H) + (4) Specific rotation: [α] _D ¹⁷ -30 ° (c0.6, CH ₃ OH) (5) Melting point: 108 ° C. (6) ¹ H NMR spectrum (400 MHz, CDCl ₃ ) δ
(ppm): 2.26 (br d, 2-H), 2.78 (dd, 2-H), 5.10 (br
d, 3-H), 3.30 (br d, 4-H), 3.60 (s, 4-OCH 3), 3.89 (br
d, 5-H), 1.79 (br t, 6-H), 1.50 (br t, 7-H), 1.64 (d
t, 7-H), 6.34 (d, 10-H), 7.38 (dd, 11-H), 6.22 (m, 12
-H), 6.22 (m, 13-H), 4.85 (ddq, 15-H), 1.29 (d, 16-
H ₃ ), 2.76 (br dd, 17-H), 9.54 (s, 18-H), 1.20 (d, 19-H
H ₃ ), 1.14 (t, 3-OCOCH ₂ C H ₃ ), 4.50 (d, 1'-H), 3.19 (dd,
2'-H), 3.45 (t , 4'-H), 1.17 (d, 6'-H 3), 2.57 (s, 3'-
N (CH ₃ ) ₂ ), 4.92 (d, 1 "-H), 1.67 (dd, 2" -Hax), 2.28 (d,
2 "-Heq), 4.72 (d, 4" -H), 4.54 (dq, 5 "-H), 1.08 (d,
6 "-H ₃ ), 1.11 (s, 7" -H ₃ ), 3.26 (s, 3 "-OCH ₃ ), 1.18 (t,
4 "-OCOCH ₂ CH ₃ )

Example 4 Compound (8) (4 "-O-depropionyl-3" -O-methylmide
Kamycin A ₃ (9-dehydro-3 "-O-methyl-3-O-propio
Nilleucomycin V)) Production method As a medium for pre-culture, the composition of starch 2.0%, glucose 1.0%, polypeptone 0.5%, wheat germ 0.6%, yeast extract 0.3%, soybean meal 0.2%, calcium carbonate 0.2% And used after adjusting to pH 7.0 before sterilization. One 100 ml Erlenmeyer flask into which 20 ml of the above-mentioned medium was dispensed was sterilized at 120 ° C. for 30 minutes, and 1.0 ml of a frozen seed of the non-spore-free defective PF1108 strain was inoculated into the Erlenmeyer flask.
Shaking culture was performed at 26 ° C for 24 hours to obtain a pre-culture solution. Then, as a medium for microbial conversion, using a composition consisting of glucose 1.8%, starch 0.9%, soybean meal 1.25%, wheat germ 0.8%, sodium chloride 0.125%, calcium carbonate 0.15%, adjusted to pH 7.0 before sterilization Used. Three 500 ml Erlenmeyer flasks into which 100 ml each of this medium was dispensed were placed at 120 ° C.
, And 1.2 ml of a methanol solution of 10 mg of compound (7) was added to each Erlenmeyer flask. Next, 5.0 ml of each of the pre-culture solutions of the PF1108 strain was inoculated, respectively, and cultured with shaking at 26 ° C. for 9 days to carry out microbial conversion. After completion of the culture, the culture was centrifuged at 3000 rpm for 10 minutes to obtain 150 ml of a transparent culture, and solids such as cells were removed. 150 ml of water was added to the solid content, and the mixture was stirred and centrifuged. The obtained washing solution was combined with the above-mentioned clear culture solution. After this was adjusted to pH 9, the converted substance was extracted twice with 300 ml each of ethyl acetate, and the ethyl acetate layer was dried over anhydrous sodium sulfate and then filtered. The filtrate was concentrated under reduced pressure, and the obtained residue was purified by preparative TLC (developing system: chloroform-methanol-concentrated aqueous ammonia (100: 10: 1)) to give compound (8) 12.0
mg was obtained. Physicochemical properties of compound (8) (1) Color and shape: colorless solid (2) Molecular formula: C ₃₉ H ₆₃ NO ₁₄ (3) Mass spectrum (EIMS): m / z 769 (M) ⁺ (4) Specific rotation Degree: [α] _D ²⁰ -27 ° (c1.0, CH ₃ OH) (5) Melting point: 121-124 ° C. (6) ¹ H NMR spectrum (400 MHz, CDCl ₃ ) δ
(ppm): 2.26 (br d, 2-H), 2.78 (dd, 2-H), 5.09 (br
d, 3-H), 3.30 (br d, 4-H), 3.60 (s, 4-OCH 3), 3.89 (br
d, 5-H), 1.78 (br t, 6-H), 1.49 (br t, 7-H), 1.64 (d
t, 7-H), 6.34 (d, 10-H), 7.38 (dd, 11-H), 6.22 (m, 12
-H), 6.22 (m, 13-H), 4.85 (ddq, 15-H), 1.29 (d, 16-
H ₃ ), 2.75 (br dd, 17-H), 9.54 (s, 18-H), 1.20 (d, 19-H
H ₃ ), 1.14 (t, 3-OCOCH ₂ C H ₃ ), 4.50 (d, 1'-H), 3.21 (dd,
2'-H), 3.45 (t, 4'-H), 3.27 (dq, 5'-H), 1.16 (d, 6'-
H ₃ ), 2.58 (s, 3'-N (CH ₃ ) ₂ ), 4.89 (d, 1 "-H), 1.57 (dd,
2 "-Hax), 2.24 (d, 2" -Heq), 3.01 (br d, 4 "-H), 4.15 (d
q, 5 "-H), 1.23 (d, 6" -H ₃ ), 1.22 (s, 7 "-H ₃ ), 3.22 (s,
3 "-OCH ₃ )

──────────────────────────────────────────────────続 き Continuing from the front page (72) Osamu Hara Inventor, Pharmaceutical Research Laboratory, Kohoku-ku, Yokohama-shi, Kanagawa Prefecture Meiji Seika Co., Ltd. Aki Nakagi, Examiner, Pharmaceutical Research Institute, Meiji Seika Co., Ltd. (58) Fields investigated (Int.Cl. ⁷ , DB name) C07H 17/08 A61K 31/7048 CA (STN) CAOLD (STN) CAPLUS (STN) REGISTRY (STN)

Claims (4)

(57) [Claims] (1) The following formula (I): [ Wherein , R ¹ is an oxygen atom , R ² is a hydrogen atom or a group of the formula COR ⁴ (where R ⁴ is a linear alkyl group having 1 to 3 carbon atoms), and R ³ is a hydrogen atom or a formula COR ⁴ Wherein R ⁴ has the same meaning as described above. Or a pharmaceutically acceptable salt thereof. 2. A compound represented by the formula (I) according to claim 1, wherein R ¹ is an oxygen atom, R ² is a propionyl group, R ³ is a hydrogen atom, or a pharmaceutically acceptable salt thereof. 3. The following general formula (II): (II) [wherein, R ² and R ³ have the same meanings as in formula (I).
You. 3-O-acyl-9- deficient in neutral sugar represented by
Dehydro-16-membered macrolide or N-oxide derivative thereof
A conductor and the following general formula (III) : (III) wherein R is an acetyl group, a propionyl group,
And a lower acyl group represented by a ril group or a valeryl group. ]
L-cladino modified at the 4-position represented by an acyl group
1-thiosugar derivative, 1-fluoro derivative or 1-
Acetoxy derivative as a starting material, 4 'Kuraisuisan
Reaction of cladinose to the group, depending on the starting material used
Deprotection step or / and reduction step of N-oxide
In the formula (I) of claim 1, R ¹ is represented by an oxygen atom, and R ² is represented by a group of the formula COR ⁴ (where R ⁴ is a linear alkyl group having 1 to 3 carbon atoms); A production method for obtaining a compound in which R ³ is represented by a hydrogen atom. 4. In the formula (I) of claim 1, R ¹ is an oxygen atom, R ² is a hydrogen atom or a group of the formula COR ⁴ (where R ⁴ has 1 to 1 carbon atoms).
A compound in which R ³ is a hydrogen atom or a group of the formula COR ⁴ (where R ⁴ has the same meaning as described above);
Or an anti-tumor agent comprising a pharmaceutically acceptable salt thereof as an active ingredient.
Fungicide .