JPS6256422A - Antibacterial agent - Google Patents

Abstract

PURPOSE:An antibacterial agent comprising diterpene as an active ingredient. CONSTITUTION:An antibacterial agent containing a diterpene shown by the formula(X is carboxyl, formly, hydroxymethyl, acyloxymethyl or alkyl; Y is hydroxyl or acyloxy; Yis acyloxy when X is carboxyl and Y is hydroxyl when X is not carboxyl) as an active ingredient. The compound shown by the formula shows improved activity inhibitory action on various Gram-positive bacteria such s Staphylococcus aureus, Bacillus subtilis, etc., and Gram-negative bacteria such s myxomycetes, etc., and is useful as an inhibitor against these bacteria. The agent is administered orally or parenterally, a dose is 0.5-50mg/kg/day, usually 1-30mg/kg/day and the agent is administered once or more times. An oral drug such as tablet, capsule, etc., or a parenteral drug such as solution, suspension, etc., may be used as a dosage form.

Description

[Detailed description of the invention] (Industrial application field) The present invention relates to antibacterial agents.

(Structure of the Invention) The present inventors are searching for physiologically active substances contained in various plants and investigating their medicinal effects. I learned that alkyl derivatives exhibit antibacterial effects.

The present invention was achieved as a result of further research based on the above findings.

To explain the present invention in detail, the diterpene which is the active ingredient of the present invention is such that X in the above formula [1] is a carboxyl group, a formyl group, a hydroxymethyl group, an acyloxymethyl group, or an alkyl group; , Y is a hydroxyl group or an acyloxy group, and when X is a carboxyl group, Y represents an acyloxy group, and when X is a formyl group, a hydroxymethyl group, an acyloxymethyl group, or an alkyl group, Y represents a hydroxyl group. Mention may be made of various diterpene compounds which represent the group. These compounds are produced as follows.

For example, compounds (compounds 1 to 4) in which X in the formula [1] is a carboxyl group and Y is an acyloxy group,
Biciferic acid in which X is a carboxyl group and Y is a hydroxyl group, for example, acetic anhydride, propionic anhydride,
It is produced by reacting an acyl with an aliphatic carboxylic acid anhydride such as butyric anhydride or valeric anhydride.

In addition, [X in formula 11 is a hydroxymethyl group, and Y
Compound 5), in which X is a methoxycarbonyl group and Y is a hydroxyl group, can be obtained by reducing methyl bicypherate, where X is a methoxycarbonyl group and Y is a hydroxyl group, with a reducing agent such as lithium aluminum hydride.

Further, a compound (compound 6) in which X is a formyl group and Y is a hydroxyl group in the formula [1] is produced by oxidizing the above compound 5 with a Jones reagent.

In addition, [in formula 11, X is an acyloxymethyl group,
Compounds in which Y is a hydroxyl group (compounds 7 to 9) are obtained by reacting the methyl bicypherate with dihydrobyran to protect the hydroxyl group with a tetrahydropyranyl group, and then
After reducing the methoxycarbonyl group to a hydroxymethyl group and then acylating with an aliphatic carboxylic acid anhydride such as acetic anhydride, propionic anhydride, butyric anhydride to convert the hydroxymethyl group to an acyloxymethyl group, Produced by hydrolyzing the tetrahydropyranyl group.

Further, in the formula [1], a compound (compound 10) in which X is a methyl group and Y is a hydroxyl group can be obtained by reacting the compound 6 with anhydrous hydrazine and then reducing it with caustic potassium.

Furthermore, in the formula [1], compounds in which X is an alkyl group other than a methyl group such as an ethyl group, a propyl group, or a butyl group, and Y is a hydroxyl group (compounds 11 to +3), the hydroxyl group of the compound 6 is replaced with tetrahydrocarbon. After protection with the pyranyl group, reaction with an alkyltriphenylphosphonium bromide or iodide (obtained from an alkyl iodide and triphenylphosphine) [Witt
ig) After reaction and removing the tetrahydropyranyl group of the resulting condensate, catalytic reduction (using Pd-C catalyst)
Manufactured by

(Effect of the invention) The diterpene represented by the formula Moe [1] is produced by Staphylococcus aureus as shown in the examples below.
aureus), Bacillus subtilis (Bacillus 5ub)
tilis) and various Durum-positive bacteria and P. tilis (Pr.
It exhibits an excellent inhibitory effect on the activity of Durham-negative bacteria such as Oteus vulgaris), and is useful as an inhibitor of these bacteria.

When using the antibacterial agent of the present invention, it is administered orally or parenterally, and the dosage is determined depending on the age, health condition, weight, etc. of the patient. Generally, the daily dose of the active ingredient is 0.5
~501 mg/kg body weight, usually 1-30 rmg/kg body weight, administered in one or more doses.

For oral administration, it is used in the form of tablets, capsules, powders, liquids, etc., and for parenteral administration, it is used in sterilized liquid forms such as liquids or suspensions.

One of these forms, solid or liquid, non-toxic carriers can be included in the composition.

As an example of a solid carrier, conventional gelatin type capsules are used. Also, the active ingredient, together with auxiliaries or alone,
It is made into tablets and packaged as a powder. These capsules, tablets and powders are generally 5-95z, preferably 25-90z! Contains active ingredients by weight. That is, for these modes of administration, 5 to 5
00 mg, preferably 25 to 250 mg of active ingredient. Water or petroleum as a liquid carrier;
Animal, vegetable, or synthetic oils such as peanut oil, soybean oil, mineral oil, and sesame oil are used.

In general, physiological saline, dextrose or similar Sho II solutions, and glycols such as ethylene glycol, propylene glycol, and polyethylene glycol are preferred as liquid carriers, and particularly in the case of injections using physiological saline, usually 0. 5-202, preferably 1-lot weight of active ingredient. In the case of liquid preparations for oral administration, the i! Liquor liquid or syrup is good. As carriers in this case, aqueous excipients such as perfumes, syrups, pharmaceutical micelles, etc. are used.

(Example) The present invention will be described in more detail below with reference to Examples.

[Antibacterial test] Agar plate dilution method shown below.
The antibacterial activity of this substance was measured using the following method.

l! First, add the sample compound to a 5% dimethyl sulfoxide aqueous solution! ! A sample reservoir [1 ml] was taken into a sterile petri dish (9 cm x 2 cm) to a predetermined concentration, and culture medium for sensitive discs (manufactured by Eiken Kagaku Co., Ltd.) 91, which had been sterilized in advance at 120°C for 15 minutes, was added to this and mixed thoroughly. It was then plated and solidified.

Using a platinum loop with an inner diameter of 1 mm+, the minimum inhibitory concentration (minimuIlinhibitory concentra- tion
centration (formerly C)) A standard strain for measurement was inoculated onto the above-mentioned plate by coating it to a length of about 2 cm, and cultured at 37°C for 18 to 20 hours. The minimum sample solution concentration at which the growth of test bacteria was completely inhibited was defined as the old C value.

The test bacteria used were those cultured at 37° C. for 18 to 20 hours using heart infusion broth medium (Eiken Chemical Co., Ltd.) and diluted 100 times with physiologically active saline immediately before use.

Using the above test method, 1 it of Staphylococcus aureus (5 taphy
l.

eoccus aureus) [FAD 209 PJ
C-1, Terajima, MS 353], dried grass [(
Sac mouth 1us 5ubtilis) [ATCC6
633 and Proteus vulgari
s) [The results of measuring the old C value of each sample compound against test bacteria such as HX-191 are shown in Tables 1 and 2 on the next page, respectively. and Prot.
For reference of activity against eus vulgaris,
The method for producing the compounds used in the above examples is illustrated below.

[Production Example] Production of Compound 4 32 mg of biciferic acid was dissolved in 0.15 ml of anhydrous pyridine, 29 μI of valeric anhydride was added, and the mixture was left at room temperature for 4 hours, then the reaction solution was poured into ice water and extracted with chloroform. did. The chloroform layer was washed successively with hydrochloric acid, 5z sodium bicarbonate, and saturated brine, dried over magnesium sulfate, and concentrated under reduced pressure. The obtained residue was subjected to preparative thin layer chromatography (n-hexane:acetone=9:I) to obtain 24 mg of Compound 4.

The specific optical rotation, 'H-NMR, and mass spectrum of this compound were as follows.

[α Kojin: + 112.97 ° (c
= 1.195. CHCl3).

'H-NMRδ', %s" 0.82 (3H+s
) r O-94 (3jl + t + J = 7-7
11z), 0.95 (3N, s), 1.1
5 (3H, d, J=6.8 Hz), 1.17 (
3Ld, J=6.8 Hz ) + 2.54 (2Lt
, J=7.7Hz), 8.89 (IH,S),
6.99 (l)l,S).

MS m/z: 400 (C,,H,,,0,,M
, 16 (relative intensity %)), 316 (M-COC,
H,,+00), 271 (M-COC,If
, -COOH,67), 201 (C,-,,0,5
), 189 (,C,?H,70,15), +75(C
,2+1. , O+I 1 >.

It should be noted that the procedure is exactly the same except that acetic anhydride, propionic anhydride, and aqueous butyric acid are used instead of valeric anhydride used in this example.
By treating each other, compounds 1 to 3 were obtained, respectively.

Preparation of Compound 5 In an argon stream, 127 mg of lithium aluminum hydride was suspended in 10 ml of anhydrous ether, and 8:51 of a solution of 276 mg of methyl bicyphelate in anhydrous ether was added dropwise at 0°C. After stirring at room temperature for 20 hours, a small amount of water was added to destroy excess reagent. The reaction solution was poured into ether, washed with saturated brine, dried over magnesium sulfate, and concentrated under reduced pressure. The obtained residue was subjected to preparative thin layer chromatography (n-hexane:acetone=
7:3) to obtain 258 mg of compound 5.

The specific optical rotation, )I-NMR and mass spectrum of this compound were as follows.

[α] +64.70” (C=0.680.
Cll, OH).

'+1-NMRδ 3: 0.89 (3)1. S)
, 0. ri3 (,3H,s)+1.20 (3H,
d, J=7. OH2)+1. ．． 21 (3H, d, J
=7. Oflz), 3.19 (1N, sep, J=
7.0 Hz), 3.49 (LH, d, J=I!, 01
1z), 3.97 (IN, d, Jll, OHz),
6.51 (br, s, OH), 6. [i9 (IH
, s ), 6.88 (IH, s ).

MS m/2: 302 (C2, H,, Ox, M
, 17,271 (M'+-CH,OH.

too), 201(C,,i,70.16),+
89 (C,,H,,0,37),+75 (C,YH
,,0,30).

Preparation of compound 6 35 mg of the above compound 5 was dissolved in acetone 11
Two drops of Jones reagent were added at 0.degree. C. and stirred for several minutes. The reaction solution was poured into saturated saline and extracted with chloroborm.

The chloroform layer was washed with saturated brine, dried over magnesium sulfate, and concentrated under reduced pressure. The obtained residue was subjected to preparative thin layer chromatography (n-hexane:acetone = 85
: 15) to obtain 22 mg of Compound 6.

The specific optical rotation, 'H-NMR, and mass spectrum of this compound were as follows.

[cx ]”: + 265.21° (c = 0.9
20. CH30)1).

IH-river δ::j-: 0.82 (3H,S), 1.
00 (3H, s), 1.20 (6H, d, J near,
0)+2), 3.16(IH, sep+J=7
．． o 1lz), 6.16 (br, s, OH), 6
．． 65 (IH, S), 6.90 (18, S), 9.8
7 (IH.

d, J=1.3112).

MS m/2: 300 (cJ, I+,, o
, 9M, I6), 271 (M-C
HO, l 00), 20! (C,,4H170,1
8), 189 (C,, ■, 70, 45),
I 75 (CJ, , 0, 40).

Preparation of Compound 7 To 5 ml of a solution of 180 mg of methylbisifunile) in straw water and methylene chloride, 200 mg of dihydrobilane and 13.6 m of pyridinium p-)luenesulfone) were added.
g was added thereto, and the mixture was stirred at room temperature for 6 hours. The reaction solution was poured into ether, washed with saturated brine, dried over magnesium oxide, and concentrated under reduced pressure.

A solution of 225 vag of the obtained residue dissolved in 31 g of anhydrous ether was added to a suspension of 83 mg of lithium aluminum hydride in anhydrous ether at 0° C. in an argon stream.
The mixture was added dropwise to nil and stirred at room temperature for 13 hours. After adding a small amount of water to decompose excess reagent, the reaction solution was poured into ether, washed with saturated brine, dried over magnesium sulfate, and concentrated under reduced pressure. The obtained residue was subjected to preparative thin layer chromatography (n-hexane:acetone: 75
: 25) to give 210 mg of the compound ([In the formula 1, X is a hydroxymethyl group and Y is a
A compound with a robilanyloxy group) was obtained.

Add 0.5 ml of pyridine to 54 mg of the compound obtained above.
After adding 0.5 ml of acetic anhydride and leaving the mixture at room temperature for 15 hours, the reaction solution was poured into ice water and extracted with chloroborm. After sequentially washing the chloroform layer with 5-dihydrochloric acid, 5' with sodium bicarbonate and saturated saline, sulfuric acid magnezi,
It was dried with a ram and concentrated under reduced pressure. Obtained residue 6011
Dissolve g in 2 ml of ethanol and dissolve pyridinium p-)
4n+g of luenesulfone) were added thereto, and the mixture was stirred for 3 hours at 55°C. The reaction solution was concentrated under reduced pressure, and the resulting residue was subjected to preparative thin layer chromatography (n-hexane:acetone = 85
: 15) to obtain 47 mg of Compound 7.

The specific rotation, H-1iMR, and mass spectrum of this compound were as follows.

[αIT)': 435-87'(C"! -1
43, CH3ON).

'H-NMR heart%': 0.95 (6H,S), 1.2
3 (6H, d, J=6.8 I2), 1.91 (3H
,s), 3.13 (IH, 5epl=6.8 Hz
), 4.16(,IH,d,J=11.Otlz)
, 4.51 (ll-1, d, J=11.o I2), 5
．． 00 (hr, s, OH), 6.67 (lH, s),
6.85 (IH,s)-MS m/z: 344 (C
,', uC'J1M, +8), 284(MC
Hyo coon.

18), 27+(M”-CH,0COCH,,+0
0), 20+(C,, 1 (720°28), +89 (
C,,H,70,44),+75(CI28I,501
72).

Compounds 8 to 9 were obtained by carrying out the same treatment except that water-collected propionic acid and butyric anhydride were used in place of the acetic anhydride used in this example.

Preparation of Compound 10 51 mg of triethylene glycol, 2.41 mg of anhydrous hydrazine, and 500 mg of hydrazine dihydrochloride were added to +40 +ag of the above compound 6, and the mixture was stirred at 140°C for 14 hours.

After cooling the reaction solution to room temperature, 2.6 g of caustic potassium granules were added and heated at 150°C for 2 hours and at 150-200°C for 2 hours.
The mixture was further stirred at 210°C for 3 hours. The reaction solution was poured into saturated brine, extracted with n-hexane, and the n-hexane layer was dried over magnesium sulfate and concentrated under reduced pressure. The obtained residue was subjected to preparative thin layer chromatography (n-hexane:acetone=9:l) to obtain 103 ff1g of Compound 10.

The specific optical rotation, 'II-NMR, and mass spectrum of this compound were as follows.

[α ko”p: + 64-00 ° (
+:"0.500.CHyOH)')l-NMR&':
3': 0.91 (3N, S), 0.93 (3H, S
), 1.16 (3H,s), 1.22 (3tl,
d, J=6.8 Hz), 1.23 (3N, d,
J=6.8Hz), 3.11 (IH, sep, J=
6.8 Hz) + 4.70 (br, s, OH) 6
．． 61 (IH,S), 6.82 (IN,s).

10MS m/z: 286 (%Ha001M, 80)
, 271 (M-CHJ, 100), 20! (C
,%H770,43), +89(C,,H,,0+8
6), +75(C,jH,,0,86).

Preparation of Compound 12 Ethyltriphenylphosphonium bromide of 84B was suspended in 1.5 ml of anhydrous tetrahydrofuran, and mixed with n-butyllithium in hexane at −25° C. under an argon atmosphere. a < 1.6 M) was added at 155 μm.

Then, after stirring at room temperature for 30 minutes, a solution of 58 mg of the compound 6 in which the hydroxyl group was protected with a tetrahydropyranyl group in 1.5 ml of anhydrous tetrahydrofuran was added.
After stirring at room temperature for 4 hours, the reaction solution was poured into saturated brine and extracted with ether. The ether extract was washed with saturated brine, dried over magnesium sulfate, and concentrated under reduced pressure.

The obtained residue was subjected to preparative J layer chromatography (n-
Hexane:acetone=98:2) to obtain 50 mg of condensate. This condensate was dissolved in ethanol of 11, and pyridinium p-)luenesulfonate was prepared.
3 rag was added and stirred at 55°C for 3 hours.

After concentrating the reaction solution, the residue was subjected to preparative chromatography (
36 mg of the compound obtained by subjecting the solution to n-hexane 7 tons (95:5) was dissolved in 1.51 g of ethyl acetate, 20 mg of Pd-C was added thereto, and the mixture was stirred at room temperature under a hydrogen stream for 13 hours. . After removing the catalyst by suction filtration, the filtrate was condensed under reduced pressure and the resulting residue was subjected to preparative thin layer chromatography (0-hexane:acetone=9:
I) to obtain 37 mg of compound I2.

The specific optical rotation, 'II-NMR, and mass spectrum of this compound were as follows.

[α Co'D: + 37-22 6
(c: I-760, CHz Otl)'H-NM
R8瘉4': 0.80 (3) 1. tlJ=6.0H
2) +0.91 (3H.

s ), 0.99 (3H, s), 1.23 (3H,
d, J=7.0 Hz), 1.24 (3N, d, J=
7.0 Hz), 3.13 (IN, sep, J=7.
0 Hz), 4.58 (br, s, OH) 6.5
4 (ILs), 6.84 (IH,s).

MS m/z: 314 (C221f, 0.M,
26), 271 (M − C,) I, , +00> ,
201(C,,H,0,14), +89(C,)I
+, 0, 24), +75(C12)11tO, 2
2) Note that Compounds 11 and 13 were obtained by treating in exactly the same manner except that methyltriphenylphosphonium bromide and pro- and rutriphenylphosphonium iodide were used instead of ethyltriphenylphosphonium bromide used in this example. Obtained.

Claims (1)

[Claims] (1) The following formula [1] ▲Mathematical formulas, chemical formulas, tables, etc.▼[1] (In the formula, X represents a carboxyl group, formyl group, hydroxymethyl group, acyloxymethyl group, or alkyl group, and Y
represents a hydroxyl group or an acyloxy group, and when X is a carboxyl group, Y represents an acyloxy group; and when X is a formyl group, a hydroxymethyl group, an acyloxymethyl group, or an alkyl group, Y represents a hydroxyl group. ) An antibacterial agent containing a diterpene as an active ingredient.