FI80688C - NY MELLANPRODUKT (8S) -8- FLUORERYTRONOLID A SOM AER ANVAENDBAR VID FRAMSTAELLNING AV HALVSYNETETISKA MACROLIDANTIBIOTIKA. - Google Patents

Description

1 80688

New intermediate (8S) -8-fluoroerytronolide A useful in the preparation of semi-synthetic macrolide antibiotics 5 Separated from application 820067

The present invention relates to (8S) -8-fluoroerytronolide A, which is useful as an intermediate in the preparation of novel macrolide antibiotics, and has the formula 10 'F'> <'CH3 CH, xVl, 0H HOn NJ L - "CH3

15 JL

° xX'oh (I CH-,

O J

Semi-synthetic macrolide antibiotics, in turn, are useful as antibacterial agents. In fact, compared to known types of erythromycins, they have similar or broader activity books and levels, are less susceptible to contamination in acidic environments, and thus allow better absorption when administered orally; their esters or salt forms are absorbed faster and more completely.

From R. A. LeMahieu et al .; J. Med. Chem. 17, 963 (1974) it is known that erythronolide A is a substrate which can be obtained from erythromycin A by selectively removing the sugars cladinose and desosamine.

It is also known that erythronolide B and 3-O-Mycarosyl erythronolide B are substrates which can be obtained directly by fermentation in appropriate amounts and thus at industrially feasible costs using erythromycin-producing microorganisms and their mutants. On the other hand, as is also well known, with the exception of the production of normal erythromycin A, these substrates have no use in other fermentations without side effects to produce antibiotics that affect erythromycin itself, whereas from an industry point of view their use seems most desirable.

It has now been found that derivatives of erythronolide A, erythronolide B or 3-O-mycarosyl erytronolide B, which can be prepared chemically in a simple and industrially feasible manner, are useful substrates for fermentation processes which: utilizing microorganisms obtained by mutation from strains and adapted for the microbiological production of erythromycin A produce new macrolide antibiotics.

The chemical process for preparing the novel intermediates of this invention is shown in the following scheme:

CU-, CH

I 3 F \ s' 3 20

L0H T - »T OH

CH3 CH - '> r I <^ CH3 C2H5 I OH 3C9H ^ OH 3 0 / c «3 0Y ^ 3 2j o 0 f11» v (III) / ^ ^ 3 0 pit | 3 F .CH3

CH, v 11 'H CH 3' Il - "0H

h ° x 3 ho n k ^ CHi ^ oH

^ s' -OH "

35 L OH I L ° H I

CH ^ "<^ CH3 CH3 ^ X ^ k ^^ CH

3 C2H5 * 0 JK0H ° 2ίΐ5 'CH3 0V> 1f CH3

O O

(I) (IV)

II

3 80688

More specifically, novel intermediates can be prepared as follows: a) Compound (I), which is erythronolide A, erythronolide B or 3-O-mycarosyl-erythronolide B, is treated with a non-aqueous acid such as glacial acetic acid or methanolic hydroxylamine hydrochloride to give ( II), namely 8,9-anhydroerytronolide A-6,9-hemiacetal, 8,9-anhydroerytronolide B-6,9-hemiacetal or 3-O-Mycarosyl-8,9-anhydroerytronolide B-6,9-hemiacetal to form 10; b) the compound (II) is reacted with a reagent capable of generating electrophilic fluorine, which is preferably a fluorooxy-perfluoroalkane (of the general formula cnF2n + i ^ F ^ or perchloryl fluoride) to form the corresponding acetal (III) in an inert organic in the presence of a solvent and at low temperature.

c) the compound (III) is reacted with an aqueous acid to form the desired compound (IV).

In step b), of the reagents of the fluorooxy-perfluoroalkanes group, the most widely used is fluorooxy-trifluoromethane, which is commercially available.

Other reagents containing fluorine atoms which have a positive charge and which can be used in this reaction include fluorooxysulfuric pentafluorine, Mole-25 alkyl fluorine, and lead tetraacetate hydrofluoric acid.

Suitable reaction solvents are chlorinated hydrocarbons such as trichloromethane (Freon 11), chloroform, methylene chloride and the like, tetrahydrofuran, and mixtures thereof. It is preferred to carry out the reaction at low temperatures, preferably between -75 and -85 ° C with continuous stirring.

4,80688

The reaction is normally complete in about 15 minutes to one hour.

It is important to emphasize that the formation of acetal (III) already involves the formation of the desired compound 5 (IV) in amounts which are not entirely negligible.

As for the third reaction step (c), organic or inorganic aqueous acids such as acetic acid or hydrochloric acid are used. Temperatures of about 30 to 150 ° C can be used for the reaction. The resulting product (IV) is recovered in pure form by recrystallization or chromatography.

compounds (II), (III) and (IV) are new.

Example 1 Preparation of 8,9-anhydroerytronolide A-6,9-hemiacetal (II).

A solution of 4.185 g (0.01 mol) of erythronolide A (I) described by R.A. LeMahieu et al. in J. Med. Chem. 17, 953 (1974), in 32 ml of glacial acetic acid, is allowed to stand for two hours at room temperature. The acetic acid is then removed in vacuo at 40 ° C and the oily residue is dissolved in 150 any chloroform. The chloroform solution is washed with saturated sodium bicarbonate solution and then with water until neutral and dried loosely with Na2SO4.

After removal of the solvent in vacuo, the crude product was purified on a silica gel column prepared from methylene chloride / methanol (1: 1).

Elution with methylene chloride / methanol 30 (98: 2) gave fractions containing only 8,9-anhydroerythronolide A-6,9-hemiacetal (II). Evaporation of these combined fractions to dryness followed by crystallization from acetone / n-hexane gave 2.750 g of compound (II) having the following characteristics:

It 5 80688 sp. 188-193 ° C + 20.5 ° (C = 1 in methanol); UV (MeOH) 210 nm (fcD6720) IR (KBr) 3630, 3520, 3495, 3400, 1710, 1465, 1450, 1440, 1415, 1400, 1370, 1350, 1310, 1285, 1230, 1200, 1170, 5 1140, 1090, 1075, 1060, 1050, 1035, 1020, 1010, 1000, 970, 950, 940, 920, 915, 905, 890, 825, 810, 800 cm-1.

Analysis for C 21 H 31 O 7 gave the following values:

Calculated (%): C 62.97 H 9.06

Found (%): C 63.12 H 9.10 Example, 2 (8S) -8-fluoroerytronolide A-6.9, 9,11-acetal (III) and (8S) -8-fluoroerytronolide A ( IV) Preparation from 8,9-anhydroerytronolide A-6,9-hemiacetal (II)

A solution of fluorooxy-trifluoromethane in CCl 4 was prepared at -80 ° C: excess (usually about twice) CF 2 OF was dissolved in CCl 4 (pre-cooled at -80 ° C with carbon dioxide ice) by slow addition of gas through a purge tube (while the cylinder containing CF 2 OF was continuously weighed on an Sartosious electric scale).

Its concentration was determined by iodometric titration.

A solution of CF 2 OF / CCl 2 F, at -80 ° C or -85 ° C, was slowly added to a solution of 4 g (0.010 mol) of 8,9-anhydroerythronolide A-6,9-hemiacetal CCl 3 In CH 2 Cl 2 (295 mL / 370 mL) at about -80 ° C, was magnetically stirred with calcium oxide (1.920 g) to remove hydrogen fluoride. The progress of the reaction was periodically monitored by high pressure liquid phase chromatography (HPLC) for the disappearance of the characteristic peak of compound (II).

After the peak of compound (II) had disappeared (or decreased), stirring was continued for five minutes and nitrogen gas was bubbled through the solution to remove excess CF 2 OF at -80 ° C and the solution was allowed to warm to room temperature. The solution was washed with a saturated solution of NaHCO 3 (650 mL) and then washed neutral with water and finally dried over Na 2 SO 4.

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Removal of the solvent gave a solid residue which was then purified by column chromatography on silica gel (1:50 ratio) prepared in methylene chloride / methanol (1: 1). Elution with increasing concentrations of methanol in methylene chloride gave fractions containing only (8S) -8-fluoroethyronolide A-6,9; 9,11-acetal (III) and fractions containing only (8S) - 8- fluoroerytronolide A (IV). After evaporation to dryness of the fractions containing compound (III) and crystallization from acetone / n-hexane, 3.435 g of product having the following characteristics were obtained: m.p. 192-3 ° C; λ max / λ + 64.7 ° (C = 1 in methanol) UV (methanol): no adsorption corresponding to the ketone group IR (KBr): 3560, 3420, 1720, 1460, 1395, 1380, 1355, 1345, 1330, 1320 , 1305, 1290, 1270, 1255, 1215, 1180 (wide), 1095, 1070, 1045, 1025, 1020, 990, 980, 970, 955, 940, 930, 910, 905, 895, 855, 840, 830, 805 cm "1

Analysis: lie gave the following values: 20 Calculated (S): C 60.27 H 8.43 F 4.54

Found (%): C 60.22 H 8.51 F 4.69

After evaporation to dryness and crystallization of the fractions containing compound (IV) from acetone / n-hexane, 145 mg of product having the following characteristics were obtained: m.p. 239-240 ° C; -3.1 ° (C = 1 in methanol) UV (methanol): 287-8 nm (£ 25.3) IR (KBr) 3610, 3550, 3480 (shoulder), 3380 (shoulder), 1735, 1700, 1460, 1405, 1390, 1380, 1350, 1325, 1300, 30 1290, 1270, 1175, 1105, 1090, 1050, 1035, 1020, 980, 960, 920, 905, 895, 875, 860 cm -1. ^ H ^ -yFOg gave the following values:

Calculated (%): C 57.78 H 8.54 F 4.35

Found (%): C 57.87 H 8.63 F 4.19

II

7 80688

Example 3 Preparation of (8S) -8-fluoroerytronolide B-6,9} 9,11-hemiacetal (III) and (8S) -8-fluoroerytronolide B (IV) from 8,9-anhydroerytronolide B-6,9-hemiacetal (II) 5 Following the same procedure as in Example 3, but starting from a solution containing 3.845 g (0.010 mol) of 8,9-anhydroerytronolide B-6,9-hemiacetal (II) (described in U.S. Patent No. 3,697,547), a crude product was obtained which, on crystallization from acetone / n-hexane, gave 3.450 g of (8S) -8-fluoro-10 erythronolide B-6.9, 9,11-acetal (III) having the following characteristics: m.p. 191-2 ° C; + 42.6 ° (C = 1.0 in methanol) UV (methanol). no adsorption corresponding to the ketone group IR (KBr): 3540, 3450, 1735, 1460, 1260, 1170, 1060, 15 1035, 980, 925, 875, 450 cm -1

Analysis for C 21 H 35 FO 6: They gave the following values: Calculated (%): C 62.67 H 8.76 F 4.72 Found (%): C 62.63 H 8.82 F 4.81

Purification of the mother liquors by silica gel column chromatography (1:50) eluting with methylene chloride / methanol (98: 2) gave 75 mg of the second product, (8S) -8-fluoroerytronolide B (IV), which had the following characteristics: mp. 247-8 °; UV (methanol): 286 nm (ε 26) 25 ° 30 ° (C = 1 in methanol) IR (KBr): 3540, 1727, 1703, 1460, 1330, 1270, 1175, 1130, 1075, 1050, 1015, 940, 920, 895, 860 cm -1

Analysis for C 21 H 37 FO 7: They gave the following values: Calculated (%): C 59.98 H 8.87 F 4.52 30 Found (%): C 59.92 H 9.00 F 4.59

Example 4 Preparation of (8S) -8-fluoroerytronolide B (IV) from (8S) -8-fluoroerytronolide B-6,9,9,11-acetal (III)

A mixture of 4.830 g (0.012 mol) of (8S) -8-fluoro-35 rierytronolide B-6.9, 9,11-acetal (III) and 3,000 ml of an aqueous solution of acetic acid (pH 3) was heated to reflux. cooling at 110 ° C with stirring for 15 minutes and then 220 ml of acetic acid was added. After one hour at 110 ° C, all starting material was dissolved. Heating was continued for half an hour and the solution was then cooled to room temperature as soon as possible, neutralized with NaHCO 3 and extracted with ethyl acetate.

After dehydration with sodium sulfate, the ethyl acetate solution was evaporated to dryness in vacuo.

After purification of the crude product by silica gel column chromatography (1:50 ratio) using methylene chloride / methanol (95: 5) as eluent, 1.8 g of product having the same chemical and physical properties as the compound isolated in Example 4 (IV) were obtained. ).

Example 5 Preparation of 3-O-mycarosyl-8,9-anhydroerytronilide B-6,9-hemiacetal (II)

A solution of 5.465 g (0.010 mol) of 3-O-mycarosyl-erythronolide B (I) described by J.R.

Martin in Biochemistry, 5, 2852 (1966), in 32 ml of 20 glacial acetic acid, is allowed to stand for two hours at room temperature.

The acetic acid is then removed in vacuo at 40 ° C and the oily residue is dissolved in 150 ml of chloroform. The chloroform solution is washed with saturated sodium bicarbonate solution and then with water until neutral and finally dried over Na2SO4. After removal of the solvent in vacuo, a crude product was purified on a silica gel column (1: 100) prepared in chloroform.

Elution with increasing concentration of methanol in chloroform gave fractions containing only 3-O-mycarosyl-8,9-anhydroerythronolide B-6,9-hemiacetal (II).

Evaporation of these combined fractions to dryness followed by crystallization from acetone / n-hexane gave 1.160 g of compound (II) having the following characteristics: li 80688 9 m.p. 85-88 ° C; Λ max δ -5 ° (C = 1 in methanol) UV (MeOH): 210 nm (ε 6850) IR (KBr): 3500, 1730, 1465, 1415, 1380, 1340, 1185, 1120, 1085, 1055, 1005 , 985, 945, 895, 810 cm -1 Analysis for C 28 H 48 ° 9 gave the following values:

Calculated (%): C 63.61 H 9.15 Found (%) s C 63.52 H 9.09

Example 6 Preparation of 3-O-mycarosyl (8S) -8-fluoroerytronolide B-6,9, 9,11-10 acetal (III) and 3-O-mycarosyl- (8S) -8-fluoroerytronolide B (IV) 3-O-Mycarosyl-8,9-anhydroerythronolide from B-6,9-hemiacetal (II)

A solution of fluorooxy-trifluoromethane was prepared in CCl 4 F at -80 ° C: excess (usually about twice) CF 3 OF was dissolved in CCl 4 F (pre-cooled to -80 ° C on dry ice) by slow addition of gas through a purge tube (while when the cylinder containing CFgOF was continuously weighed with a Sartorious electric balance).

Its concentration was determined by iodometric titration.

A solution of CFgOF / CClgF, at -80 ° C or -85 ° C, was slowly added to a solution of 5.285 g (0.010 mol) of 3-O-mycarosyl-8,9-anhydroerythronolide β-6,9-hemiacetate. (II) in CCl 3 F / CH 2 Cl 2 (295 mL / 370 mL) at about -80 ° C, was magnetically stirred with calcium oxide (1.920 g) to remove hydrogen fluoride.

k

The progress of the reaction was monitored periodically by high pressure liquid chromatography (HPLC) for the disappearance of the peak characteristic of compound (II).

After the peak (or decrease) of compound (II) had disappeared (or decreased), stirring was continued for another five minutes and nitrogen gas was bubbled through the solution to remove excess CF 2 OF at -80 ° C and the solution was allowed to warm to room temperature. The solution was washed with saturated 35 · NaHCO 3 solution (650 mL) and then washed to nitrate with water and finally dried over Na 2 SO 4. Removal of the solvent gave a solid residue which was then purified by silica gel column chromatography (1: 100) prepared in chloroform.

Elution with increasing concentration of methanol in chloroform gave fractions containing only 3-O-O-mycarosyl- (8S) -8-fluoroerytronolide B-6,9 * 9,11-acetal (III) and fractions containing only 3-0 -mycarosyl- (8S) -8-fluoroerytronolide B (IV).

After evaporation to dryness of the fractions containing compound (III) and crystallization from acetone 10, 1.100 g of material having the following characteristics were obtained: m.p. 175-7 ° C; -9.8 ° (C = 1 in methanol) UV (methanol): no adsorption corresponding to the ketone group IR (KBr): 3540, 3490, 1720, 1460, 1395, 1380, 1365, 1355, 1330, 1320, 1290, 1275, 1265 , 1245, 1205, 1185, 1165, 15 1145, 1120, 1100, 1080, 1060, 1045, 1015, 1005, 990, 960, 940, 925, 915, 900, 880, 865, 840, 815 cm -1

Analysis for C 28 H 28 FO 3 gave the following values: Calculated (%): C 61.52 H 8.67 F 3.47 Found (%): C 61.46 H 8.51 F 3.48 20 Fractions containing After evaporation of compound (IV) to dryness and crystallization from acetone / n-hexane, 290 mg of product having the following characteristics were obtained: m.p. 214-5 ° C; -73.2 ° (C = 1 in methanol) 25 UV (methanol) 287 nm (£ 25.3) IR (KBr): 3570, 3550, 3460 (shoulder), 1745, 1735, 14.65, 1380, 1365, 1340, 1300, 1275, 1250, 1180, 1115, 1080, 1055, 1010, 1000 (shoulder), 985, 945, 935, 920, 895, 855, 840, 810 cm -1 Analysis for C 29 H 49 FO 10 gave the following values:

Calculated (%): C 59.55 H 8.75 F 3.37 Found (%): C 59.68 H 8.60 F 3.48

Example 7 Preparation of 3-O-mycarosyl- (8S) -8-fluoroerytronolide B (IV) 35 from 3-O-mycarosyl- (8S) -8-fluoroerytronolide B-6,9,9,11-acetal (III) 11 n 80688

A mixture of 6.560 g (0.012 mol) of 3-O-mycarosyl- (8S) -8-fluoroerytronolide B-6.9, 9.11-acetal (III) and 3,000 ml of aqueous acetic acid (pH 3), was kept at room temperature with stirring until complete dissolution and then 220 ml of acetic acid were added.

The solution was allowed to stir at room temperature until the starting compound (III) was completely disappeared (checked by high pressure liquid chromatography), neutralized with NaHCO 3 and extracted with ethyl acetate.

After removal of water with sodium sulfate, the ethyl acetate solution was evaporated to dryness in vacuo.

Purification of the crude product by silica gel column chromatography (1:50 ratio) methylene chloride / methanol (95: 5) as eluent gave a product having the same chemical and physical properties as the compound (IV) isolated in Example 7.

Example 8 Preparation of (8S) -8-fluoroerytronolide A (IV) from (8S) -8-20 fluoroerytronolide A-6,9} 9,11-acetal (III)

A mixture of 5.020 g (0.012 mol) of (8S) -8-fluoroerytronolide A-6,9} 9,11-acetal (III) and 3,000 ml of aqueous acetic acid (pH 3) was heated to reflux at 110 ° C: with stirring for 15 minutes and then 220 ml of acetic acid were added.

After one hour at 110 ° C, all starting material was dissolved.

Heating was continued for half an hour and the solution was then cooled to room temperature as quickly as possible, neutralized with NaHCO 3 and extracted with acetic acid.

After removal of water with sodium sulfate, the ethyl acetate solution was evaporated to dryness in vacuo.

The crude solid was then purified by silica gel 35 column chromatography (1:50) prepared in methylene chloride.

12 80688

Elution with increasing concentration of methanol in methylene chloride gave fractions still containing the starting compound, (8S) -8-fluoroerytronolide A-6,9, 9,11-acetal (III) and fractions containing only (8S) -8- 5 f fluoroerythrolide A (IV)

Repeating the reaction with the recovered starting compound followed by chromatographic purification gave fractions containing (8S) -8-fluoroerytronolide A. Evaporation of all the combined fractions to dryness and then recrystallization of the precipitate gave 0.725 g of (8S) -8-fluoroerytronolide A (IV) having the same properties as in Example 3.

Il

Claims (4)

13 80688 Patent Tivatimus (8S) -8-Fluoro-Rhytronolidi A, yoka on the caudal tuberculosis velituotteena valmistettaessa uusia macrolidianti-biootteja, yes young kaava on 5 o F> <^ CH3 ch ,, 11 -0H HO, 3 NJ L ^ CH3 ^^ OH 10 X ° h X CHl ^ fv CH? 3 (2S) -8-fluoroerythrolide A, which is useful as an intermediate in the preparation of new macrolide antibiotics and which has the formula 0 P> r '' CH3 '' (l SOH HOv N b-C1,3 30 ^ -OH I ^ OH I CH3 ^ f \ CH, C2H5 'o i ^ OH H CH 35 O 3