Method of 4"-epi-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycine a preparation

A process for the preparation of 4'-epi-9a-methyl-9a-aza-9a-hom'Oerythromycin A or a pharmaceutically acceptable salt thereof, characterized in that methylation of 4'-epi-9-deoxo-9a-aza- 9α-homoerythromycin A formaldehyde in the presence of a reducing agent selected from the group consisting of formic acid, sodium cyanoborohydride or hydrogen and a noble metal catalyst, in an inert solvent at a temperature of 20 to 100 ° C.

The compound of the invention and its pharmaceutically acceptable salts can be used as an antibacterial agent.

The present invention relates to the antibacterial 4'-epi-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin A, its pharmaceutically acceptable salts and intermediates useful in the preparation of erythromycin A.

Erythromycin A is a well known macrolide antibiotic of Formula I which has found significant clinical use.

<img file="CS241069B2_D0001.tif" />

The therapeutically useful compound is the 4'-epimer of the previously reported erythromycin A of formula II (R = methyl and III (R = hydrogen)]

<img file="CS241069B2_D0002.tif" />

which are the subject of Belgian patent No. 892,357. In the Belgian patent, the compound of formula II is called the N-methyl derivative "11-aza-10-deoxo-10-dihydroerythromycin A" by the name previously used by Kobrehel et al., U.S. Patent 4,328,334; for the precursor of this compound of formula III. Since erythromycin A derivatives are ring extended (homo), nitrogen is replaced by carbon (aza), we prefer to name 9-deoxo-9a-aza-9a-homoerythromycin A. This compound can also be named as a 10-aza-14-hexadecanolide derivative.

Certain of the new intermediates are also

4'-epimers of the foregoing known compounds. Thus, the 4'-epi-9-deoxo-9a-aza-9a-homoerythromycin A is the 4'-epimer of the above compound of formula III and the 4'-epierythromycin A oxime is the 4'-epimer of erythromycin A oxime described by Djokic et al. No. 3,478,014.

The present invention provides a process for the preparation of 4'-epi-9-deoxe-9a-methyl-9a-aza-9a-homoerythromycin A of formula IV or pharmaceutically acceptable salts thereof, characterized in that it is methylated with 4'-epi-9- deoxo-9α-aza-9α-homoerythromycin A formaldehyde in the presence of a reducing agent selected from the group consisting of formic acid, sodium cyanoborohydride or hydrogen, and a noble metal catalyst, in an inert solvent at 20 to 100 ° C.

<img file="CS241069B2_D0003.tif" />

(IV) R = methyl, Z = Z¹ = hydrogen, (V) R = hydrogen, Z and Z¹ together form an oxygen atom, (VI) R = Z = Z¹ = hydrogen atom.

The therapeutically active compound (IV) of the present invention exhibits a relatively broad spectrum of antibacterial activity that includes erythromycin A-sensitive strains and moreover fully includes the major respiratory pathogen Homophilus influenzae. It exhibits high oral absorption and an excellent half-life in vivo, making Compound IV particularly valuable for treating bacterial infections in mammals.

The intermediates for the synthesis of 4'-epi-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin A (IV) are:

a) A compound selected from the group consisting of 4'-epi-9a-aza-9a-homoerythromycin A and its 9-deoxo derivative of formulas V and VI above.

b) 4'-epierythromycin A oxime.

c) A compound selected from the group consisting of 9α-benzyloxycarbonyl-9-deoxo-4'-deoxy-4'-oxo-9α-aza-9α-homoerythromycin A of formula VII; 9-deoxo-4'-deoxy-4'-oxo-9a-methyl-9a-aza-9a-homoerythromycin A of formula (VIIa) and their corresponding 2'-O-C 2 -C 3 alkanoyl derivatives of formulas VIII and VIIIa . Acetyl is preferably used as the 2'-O-alkanoyl derivative having 2 to 3 carbon atoms.

<img file="CS241069B2_D0004.tif" />

<img file="CS241069B2_D0005.tif" />

(VII) R¹ - benzyloxycarbonyl, R² - H (VIII) R 1 = benzyloxycarbonyl, R 2 = - (C 2 -C 3) alkanoyl (VIIa) R 1 = methyl, R 2 = H (Fairy) R 1 = methyl, R 2 -alkanoyl of 2 to 3 C atoms

d) A compound selected from the group consisting of 2'-O-acetyl- and 2'-O-propionyl-9-deoxo-9a-benzoyloxycarbonyl-9a-aza-9a-homoerythromycin A of formula IX. Particularly valuable is the 2'-O-acetylderivative of formula IX

<img file="CS241069B2_D0006.tif" />

(X) R 3 = hydroxyl, (XI) R 3 -methyl.

The antibacterial compound of the present invention, 4'-epi-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin-A (IV), is readily prepared in a number of ways from erythromycin A. new - and known compounds as intermediates include the following transformations:

<img file="CS241069B2_D0007.tif" />

(A) epimerization to C-4 ', (B) ring expansion to introduce a nitrogen atom to the 9a position, (C) removal of the 9-oxo group, (D) methylation of 9a-N-, together with any optional - introduction or - by cleavage of protecting groups. Preferred are the following transformations performed in one or - i - other sequences:

(A) (B) (C) (D), (B) (A) (C) (D), or (B) (C) (D) (A).

(IX) R 1 = benzyloxycarbonyl, R 2 = alkanoyl of 2 to 3 C atoms.

e) Compound from the group consisting of - 4'-epi-9-deoxo-9a-hydroxy-9a-aza-9a-homoerythromycin A 3'-N-oxide and 4'-epi-9-deoxo-9a-methyl-9a- aza-9α-homoerythromycin A 3'-N-oxide of formulas (X) and (XI).

The various intermediates and end products are isolated by standard methods - (for example, extraction, precipitation, evaporation, chromatography, crystallization).

(A) (B) (C) (D)

The operating sequence (A) (B) (C) (D) first involves the conversion of erythromicin A - (I) to

4-epierythromycin A according to the method of Sciavolino et al. (Supra). The 4-epi-erythromycin A is then converted in almost quantitative yield to the 4-epi-erythromycin A oxime by reaction with a hydroxylamine or preferably a hydroxylammonium salt, such as a hydrochloride. Under the preferred conditions found now, at least one molar equivalent, usually an excess, for example 10 to 30 equivalents of hydroxylamine, is used in an excess of a weakly basic tertiary amine (preferably p- pyridine) as a solvent at a temperature of 0 to 50 ° C, preferably at rooms.

Incurred 4^at-epi-erythromycin oxime is rearranged to the 4'-epi-9a-aza-9a-homoderivative (V) by Beckman rearrangement. Preferred conditions use an excess (e.g., 3-4 molar equivalents) of an organic sulfonyl chloride, preferably methanesulfonyl chloride, which is reacted with oxime (in the free base or acid salt form) in a mixture of a lower ketone (e.g., methyl ethyl ketone, acetone) and water, containing a large excess of sodium bicarbonate at a temperature of from 0 to 50 ° C, preferably from 0 to 30 degrees Celsius.

The amide carbonyl at C-9 of the compound of formula (V) is then preferably reduced to the corresponding dihydroderivative, for example 4'-epi-9-deoxo-9a-aza-9a-homoerythromycin A (VI) with sodium borohydride (preferably using an excess to force the reaction to proceed in full time, but at least using two equivalents). The reduction is carried out in a suitable protic solvent such as a lower alkanol (preferably methanol) at 0 to 50 ° C (preferably at or below 38 ° C). Excess NaBH 4 was carefully quenched by addition of dilute aqueous acid to the reaction mixture.

The final methylation to give compound IV is carried out by reductive methylation using formaldehyde in the presence of a reducing agent such as hydrogen and a noble metal catalyst, sodium cyanoborohydride, or preferably formic acid. The reaction is preferably carried out with at least one equivalent of formaldehyde and one equivalent of formic acid in an inert solvent at 20 to 100 ° C. The preferred solvent is chloroform. In this solvent, the refining components are preferably mixed at room temperature and then heated to reflux to complete the reaction.

Alternatively, methylation of a compound of formula (VI) to (IV) is accomplished by oxidative protection of the dimethylamino group to the corresponding N-oxide (simultaneously forming the 9α-N-hydroxy derivative), methylating with methyl iodide with concomitant (at least partially) 9α-N-deoxygenation methyl 3'-oxide. The oxidation of the compound of formula VI is readily accomplished by reaction with hydrogen peroxide, generally in an excess of at least two molar equivalents necessary in an organic solvent inert to the reaction at a temperature of 10 to 50 ° C, preferably at room temperature. In this way, 9α-hydroxy-3'-N-oxide (X) is formed. This compound is methylated and deoxygenated to a compound of formula (XI) with methyl iodide, preferably in an organic solvent inert to the reaction, for example methylene chloride at 0 to 50 ° C (preferably room temperature), preferably in the presence of a solvent insoluble base which neutralizes an acid formed (e.g. HJ when methyl iodide is used as the methylating agent). With methylene chloride as the solvent, the preferred base is potassium carbonate. Thus, excess base and potassium iodide formed are completely removed by simple filtration prior to isolation of 9α-methyl-3'-N-oxide (XI). Finally, removal of the 3'-N-oxide group is readily accomplished by hydrogenation on a noble metal catalyst or Raney nickel. In this hydrogenation, temperature and pressure are not critical, for example, the temperature may be in the range of 0 to 100 ° C and the pressure in the range of less than about 1 to 10 MPa or more. Most preferred is room temperature and a slight overpressure, for example 0.2 to 0.8 MPa. Suitable noble metal catalysts are palladium, rhodium and platinum, supported or unsupported on supports known in the art of catalytic hydrogenation. Preferred catalysts are palladium on carbon and Raney nickel.

(A) (C) (D)

Reaction sequence (B) (A) [C) (D) involves first converting erythromycin A (I) to 9-deoxo-9α-aza-9α-homoerythromycin (III) via erythromycin A oxime and 9α-aza-9α-homoerythromycin, by the method of Kobrehel et al. (supra). In this context, the new procedure described above for 4^atThe epi-erythromycin A-oxime is preferably used to prepare the intermediate erythromycin A-oxime.

The 2'-hydroxy group of compound (III) is first protected in the form of its acetate or propionate. The acylation is selectively carried out by reacting compound III with a limited excess of acetic or propionic anhydride in an inert solvent (e.g. methylene chloride) at 0 to 30 ° C (preferably at room temperature). A limited excess of anhydride is used to compensate for the reagent consumed in the side reactions, for example unwanted acylation of other groups, especially 9α-nitrogen.

The resulting 2'-alkanoyl derivative having 2 to 3 carbon atoms is then protected on the 9α-nitrogen with a benzyloxycarbonyl group. This compound IX is formed by reacting the above 2'-ester with carbobenzoxy chloride in a reaction inert solvent in the presence of a base. Particularly suitable are Schotten-Baumann conditions, for example the reaction of the 2'-ester with the acid chloride is carried out under suitable alkaline conditions, for example in aqueous tetrahydrofuran, while maintaining the pH at 7.5 to 7.5.

8.5 with dilute sodium hydroxide during the addition of the acid chloride and during the reaction. The temperature is not critical, but generally ranges from 0 to 50 ° C, preferably at room temperature.

The C-4 &amp; apos; -hydroxyl in compound IX is then oxidized to the C-4 &amp; apos; -oxo compound VIII by treatment with a mixture of oxalyl chloride and dimethyl sulfoxide at low temperature (-40 to -80 [deg.] C.) in an organic reaction inert solvent (e.g. methylene chloride). the reaction mixture is treated with an excess of a tertiary amine (e.g. triethylamine) in the cold. The alkanate protecting group is then removed by solvolysis, preferably by contacting an excess of methanol at a temperature of 0 to 100 ° C to give compound (VIII).

Raney-nickel hydrogenation using the conditions described above converts compound VII to 4'-epi-9-deoxo-9α-aza-9α-homoerythromycin A (VI). This compound is then converted to the 9α-N-methyl derivative (IV) according to one of the alternative methods described above.

(B) (C) (D) (A)

This reaction sequence involves first converting erythromycin A to the above compound of formula II using the methods detailed in the "Preparations" section below. The C-4-epimerization is then carried out by the procedures and methods described above. The 2 &amp; apos; -hydroxy group is protected by acylation, the 4 &amp; apos; -hydroxy group is oxidized to the 4 &amp; apos; -oxo group, preferably the substituted trifluoroacetic anhydride and oxoxyl chloride. The protecting acyl group is removed and the 4'-oxo group is catalytically hydrogenated to the desired 4'-dimer hydroxy group. In this case, the preferred catalyst is Raney nickel.

Since the compound IV of the present invention contains two basic nitrogen atoms, pharmaceutically acceptable mono-acid addition salts of the free base IV with the acid are formed depending on whether one equivalent of acid or at least two equivalents of acid is used. Salts are generally formed by combining reagents in a reaction-inert solvent, and if the salt does not precipitate directly, it is isolated by concentration and / or addition of a non-solvent. Suitable pharmaceutically acceptable addition salts are compounds with HCl, HBr, HNO3, H2SO4, HO2CCH2CH2CO2H, cis- and trans-HOC3C'HCHCO2H, CH3SO3H, and p-CH3C6H4SO3H, although the salts are not limited to these acids.

The antibacterial activity of the compounds of formula IV is demonstrated by measuring the minimum inhibitory concentration (MIC) in₍₍tg / ml for various microorganisms in brain-cardiac infusion medium. Generally, a two-fold dilution of the test compound is used, with an initial test compound concentration ranging from 50 to 200 (tg / ml). The sensitivity (MIC) of the test organism is the lowest compound concentration capable of producing complete growth inhibition observed with the naked eye. -9-deoxo-9α-methyl-9α-aza-9α-homoerythromycin A IV with control erythromycin A is shown in Table I.

AND

Table I

In vitro activity of compound (IV)

<td colspan="3">repeated MIC values</td>

<td>den 1</td><td>den 2</td><td></td>

<td>А В</td><td>AND</td><td>В</td>

<td rowspan="2">Staph. aur.</td><td> 005</td><td> 0,05</td><td> 0,20</td><td> 0.05</td><td> 0,39</td>

<td> 052</td><td> 0,10</td><td> 0,20</td><td> 0,10</td><td> 0,39</td>

<td></td><td> 400</td><td> 3,12</td><td> 3,12</td><td> 6,25</td><td> 12,5</td>

<td>Stapli. epi.</td><td> 111</td><td> 0,05</td><td> 0,10</td><td> 0,05</td><td> 0,20</td>

<td>Shard, faec.</td><td> 006</td><td> 0,78</td><td> 1,56</td><td> 0,78</td><td> 0,78</td>

<td>Shard, pyog.</td><td> 203</td><td> 0,025</td><td> 0,025</td><td> 0,025</td><td> 0,025</td>

<td>Shard, pneumo.</td><td> 012</td><td> 0,025</td><td> 0,025</td><td> 0,025</td><td> 0,025</td>

<td>E. coli</td><td> 125</td><td>(and)</td><td> 6,25</td><td>(and)</td><td> 6,25</td>

<td></td><td> 129</td><td>(and)</td><td> 1,56</td><td>(and)</td><td> 6,25</td>

<td></td><td> 266</td><td>(and)</td><td> 3,12</td><td>(and)</td><td> 6,25</td>

<td></td><td> 470</td><td> 3,12</td><td> 0,78</td><td> 3,12</td><td> 0,78</td>

<td rowspan="2">Kleb. Am.</td><td> 009</td><td>(and)</td><td> 12,5</td><td>(and)</td><td> 12,5</td>

<td> 031</td><td>(and)</td><td> 12,5</td><td>(and)</td><td> 12,5</td>

<td>Kleb. oxy.</td><td> 024</td><td>(and)</td><td> 12,5</td><td>(and)</td><td> 12,5</td>

<td>Trap. mult.</td><td> 001</td><td> 1,56</td><td> 0,10</td><td> 1,56</td><td> 0,10</td>

<td>Serr. mař.</td><td> 017</td><td>(and)</td><td> 50</td><td>(and]</td><td> 50</td>

<td>Neiss. sic.</td><td> 000</td><td> 1,56</td><td> 0,20</td><td> 3,12</td><td> 0,39</td>

<td>Ent. aerog.</td><td> 040</td><td>(and)</td><td> 12,5</td><td>(and)</td><td> 12,5</td>

<td>Ent. cloac.</td><td> 009</td><td>(and)</td><td> 25</td><td>(and)</td><td> 25</td>

<td>Prov. strua.</td><td> 013</td><td>(and)</td><td> 50</td><td>(and)</td><td> 50</td>

<td>H. influ.</td><td> 012</td><td> 3,12</td><td> 0,39</td><td> 1,56</td><td> 0,39</td>

<td></td><td> 036</td><td> 6,25</td><td> 0,39</td><td> 3,12</td><td> 0,39</td>

<td></td><td> 038</td><td> 6,25</td><td> 0,39</td><td> 3,12</td><td> 0,78</td>

<td></td><td> 0,42</td><td> 1,56</td><td> 0,39</td><td> 1,56</td><td> 0,39</td>

<td></td><td> 051</td><td> 3,12</td><td> 0,39</td><td> 3,12</td><td> 0,78</td>

<td></td><td> 073</td><td> 3,12</td><td> 0,39</td><td> 3,12</td><td> 0,78</td>

<td></td><td> 078</td><td> 1,56</td><td> 0,39</td><td> 1,56</td><td> 0,39</td>

<td></td><td> 081</td><td> 3,12</td><td> 0,39</td><td> 3,12</td><td> 0,78</td>

(a) greater than 50

Erythromycin A control В compound (IV)

In addition, Compound IV was tested in vivo by a well-known mouse protection assay or a microbiological assay to determine serum levels in various mammals (e.g., mice, rats, dogs). Using rats as the test species, compound IV is well absorbed at oral dosing, yielding high and long-lasting serum levels.

For the treatment of systemic infections in mammals, including humans, caused by susceptible microorganisms, Compound IV is dosed in an amount of 2.5 to 100 mg / kg / day, in divided doses or preferably in a single dose per day. Variations in dosage depend on the individual being treated and the sensitivity of the microorganism. These compounds are dosed orally or parenterally, preferably orally. The susceptibility of a microorganism isolated in a hospital is routinely tested in clinical laboratories by well known plate methods. Compound IV is generally a preferred compound as seen from the relatively large inhibitory zone for bacteria causing therapeutic infections.

The preparation of optimal dosage forms is carried out by conventional methods of pharmaceutical practice. For oral administration, the compounds are formulated either alone or in combination with pharmaceutical carriers such as inert solid diluents, aqueous solutions or non-toxic solvents in dosage forms such as gelatin capsules, tablets, powders, cachets, syrups and the like. Such carriers include water, ethanol, benzyl alcohol, glycerin, propylene glycol, vegetable oils, lactose, starch, talc, gelatin, gums, and other well known carriers. Parenteral dosage forms for the above systemic use require that they be dissolved or suspended in a pharmaceutically acceptable carrier such as water, sodium chloride solution, sesame oil, and the like. Agents which improve the suspensibility and dispersion quality of the parenteral form may also be added.

For the topical treatment of infections in animals, including humans, caused by susceptible microorganisms, compound IV is formulated by methods well known in the pharmaceutical arts for lotions, ointments, creams, gels and the like in concentrations ranging from 5 to 200 mg / cm³dosage forms, preferably in the range of 10 to 10 mg

14 up to 100 mg / cm³. The dosage form is applied to the site of infection generally at least once a day.

The present invention is illustrated by the following examples. However, it is to be understood that the invention is not limited to certain details of these examples. Unless otherwise stated, all operations are carried out at room temperature, all solvents are evaporated under a bath vacuum of 40 ° C or less, all temperatures are in degrees Celsius, all thin layer chromatography is performed on commercial silica gel plates [using the eluent indicated in parentheses), all solvent ratios are by volume.

Example 1

4'-epi-erythromycin A oxime [Oxime of 4'-epimer (I)]

4'-epi-Erythromycin A (50 g, 0.0646 mel) was dissolved in 265 ml pyridine. Hydroxylamine hydrochloride (112.2 g, 1.615 mol) was added and the suspension was stirred for 16 hours. The reaction mixture was evaporated to a thick slurry, diluted with 300 mL of isopropanol, stirred well, filtered and washed with 3 X 100 mL of isopropanel. The filtrate and washings were combined, evaporated to a water soluble foam, triturated in ether to give the crude title compound as the hydrochloride (100 g). This was purified by shaking between CH 2 Cl 2 and defective sodium bicarbonate, adjusted to pH 9.5 with dilute sodium hydroxide. The aqueous phase was separated, washed with ethyl acetate and then with ether. All organic phases were combined, dried over sodium sulfate and evaporated to give the title compound as a white foam, 59.5 g, Rf 0.5 (60: 10: 1 CHCl 2):

: СНзОН: Conc. NH 4 OH);

WMR (CDCI3)

2.31 (6H, s, (CH3) 2N-),

3.32 (3H, s, cladinose CH3O-).

Example 2

4 '-epi-9α-aza-9α-homoerythromycin A (IV)

The product prepared in the previous example (59.2 g, 0.0787 mol) was dissolved in 400 ml of acetone. Add NaHCO 3 suspension (60 g) in 225 mL water. Methanesulfonylchloride (36.3 g, 24.5 mL) in 50 mL acetone was then added portionwise over 10 minutes while maintaining the temperature at 30 ° C with a cooling bath. The mixture was stirred for 4.5 hours, the acetone was evaporated, dichloromethane (400 ml) was added to the residue, and the pH was adjusted to 5.6 with 6N hydrochloric acid. The aqueous phase was separated, washed with two additional portions of dichloromethane and adjusted to pH 9.5 with 6N NaOH. The alkaline solution was extracted twice with fresh dichloromethane, once with ethyl acetate and once with ether. The alkaline organic extracts were combined, dried over sodium sulfate and evaporated to give the title compound as a foam, yield 41 g, thin layer chromatography, Rf 0.4 (60: 10: 1. CH 2 Cl 2: NH 2: conc. NFLiOH);

- 1 H NMR (CDCl 3) δ:

2.27 (6H, s, / CH 5/2 N -),

3.29 (3H, s, cladinose СШ0), ¹³CNMR (COCh, / CH3 / 4Si internal standard) PPm

177.24 (lactone C = 0),

163.53 (amide C = O),

102.29 and 95.24 (C-3, C-5),

40.22 ((CH 3) 2 N -).

Example 3

2'-O-acetyl-9-deoxo-9a-aza-9a-homoerythrG '' mycin A [2'-O-acetate (III)]

9-deoxo-9α-aza-9α-homoerythromycin A [10 g, 0.0136 mol; (III), U.S. Pat. No. 4,328,334) is dissolved in 150 ml of dichloromethane. Acetic anhydride [1.39 g, 1.28 mL, 0.0136 min) was added and the mixture was stirred for 3 hours. Acetylation was monitored by thin layer chromatography and 0.25 ml acetic anhydride and then 0.5 ml acetic anhydride were added to complete the reaction, while stirring the reaction mixture first

1.5 hours and then 1 hour. The reaction mixture was diluted with water and adjusted to pH 11 with dilute sodium hydroxide solution. The organic phase was evaporated to give 11.5 g of a foam. The foamed product (10 g) was chromatographed on 300 g of silica gel using 9: 1 CH 2 Cl 2: NH 2 O as the eluent for monitoring. The less polar impurities (3.6 g) eluted first and then the purified title product which was isolated as a foam (2 g), thin layer chromatography, Rf 0.2 (90: 10: 1, CH 2 Cl 2: СНзОН: conc. NHiOH);

HINMR (CDCl 3) δ:

O

2.02 [3H, s, C-2 ', -O-C (R)),

2.26 (6H, s, (CH3) nN-J,

3.35 (3H, s, cladinose CH3O-).

By the same method, substituting propionic anhydride for acetylanhydride, the corresponding 2-O-propionylderivative is prepared.

Example 4

2'-O-acetyl-9-deoxo-9α-benzyloxycarbonyl-9α-aza-9α-homoerythromycin A [(IX), R² = acetyl]

The product prepared in the previous example (1.7 g, 0.00219 mol) was dissolved in 70 ml of a 5: 2 mixture of tetrahydrofuran and water. The pH was adjusted to 8 with dilute sodium hydroxide. Carbobenzoxychloride (0.51 g, 0.427 ml, 0.003 mol) and the mixture was stirred for 2 hours while adding additional dilute sodium hydroxide to maintain the pH at 8. Since the reaction was incomplete according to thin layer chromatography, additional carbobenzoxy chloride (0.3 mL) was added and the reaction continued for an additional 3 hours while maintaining the pH at 8. The reaction mixture was washed with water and ethyl acetate, adjusted to pH 9, 6 and the aqueous phase was washed with dichloromethane. The organic phases were combined, dried over sodium sulfate and evaporated. 2.4 g of a foam are obtained which is chromatographed on 85 g of silica gel, eluting with 170: 101: 1, CH3Cl2: N2O2: conc. NH4OI-I. The pure fractions were combined, evaporated to a foam, taken up in dichloromethane and concentrated until the product crystallized. 1.2 g, mp 122 ° C, thin layer chromatography Rf 0.4 (90: 10: 1, CH 2 Cl 2: NH 2 OH: conc. NH 4 OH);

iHNMR (CDCl3) δ:

O

II

2.00 (3H, s, C-2 ', —O — С — СНз),

2.27 (6H, s, (CH3) 2N-),

3.35 (3H, s, cladinose CH3O-J, ¹³CNMR (CDCh, / CH5 / 4Si internal standard) ppm:

157.10 (carbamate C = O),

137.0, 127.55 and 127.92 (aromatic ring),

40.6 (/ CH + N-).

In the same manner, the 2'-O-propionyl derivative of the previous example is converted to the corresponding 2'-O-propionyl-9α-benzyloxycarbonyl derivative.

Example 5 2'-O-acetyl-9a-benzyloxycarbonyl-9-deoxo-4'-deoxo-4'-oxo-9a-aza-9a-homoerythromycin A [(VIII), R2 = acetyl]

Oxalyl chloride (4.37 g, 3.0 mL, 0.0344 mol) was dissolved in 25 mL of dichloromethane and cooled to 60 ° C. Add dimethylsulfoxide (6.70 g, 6.09 mL, 0.0856 mol) in 9 mL of dichloromethane. The reaction mixture was maintained at -60 ° C for 10 minutes and at this temperature the product of the previous example (5.2 g, 0.00572 mol) in 16 ml of dichloromethane was added. After an additional 25 minutes at -60 ° C, triethylamine (17.3 g, 23.9 mL, 0.172 mol) was added and the mixture was allowed to warm to room temperature, diluted with 50 mL of water and excess sodium bicarbonate. The organic phase was separated, dried over sodium sulfate and evaporated to give the title product as a sticky foam. Yield 6.8 g, thin layer chromatography Rf-0.6 (90: 10: 1, CH 2 Cl 2: NH 4 OH: conc. NH 3 OH);

• HNMR (CDCh):

O

II

2.05 (3H, s, C-2 — O — С — СНз),

2.25 (6H, s, (CH3) 2N-),

3.32 (3H, s, cladinose СНзО-],

7.37 (5H, s, aromatic protons);

MS: major peaks at m / e 53G and 518 [Nbenzyloxycarbonyl ion and glycone (minus both sugars by Cl ', C-5 cleavage)], 200 (base peak, desosamine-derived fragment), 125, (neutral sugar-derived fragment) ). This intermediate is preferably used immediately in the next step.

In the same manner, the corresponding 2'-O-propionyl-4'-oxo derivative was prepared from the 2'-O-4-propionyl compound of the preceding example.

Example 6

9a-Benzyloxycarbonyl-9-deoxo-4'-deoxy-4 '^and-oxo-9α-aza-9α-homoerythromycin А (VII)

The product prepared according to the preceding example, 1.0 g, was stirred for 65 hours in 25 ml of methanol and then evaporated to a foam. The foam was dissolved in dichloromethane, washed with saturated sodium bicarbonate solution and re-evaporated to a second foam. The second foam is chromatographed on 20 g of silica gel using a 13: 1 mixture of CH 2 Cl 2: NOSO as eluent. Fractions containing pure product were combined and evaporated. The title compound is obtained. in the title in the form of foam. Yield 336 mg, thin layer chromatography, Rf 0.4 (9C: 10: 1, CH 2 Cl 2: NH 2 OH: conc. NH 4 OH;

¹³CNMR (CDCh, / CH3 / 4SI internal standard) ppm:

210.87 (C-4 ', C = O)

176.03 (lactone C = O),

157.41 (carbamate C = 0),

136.31, 128.2 and 128.0 (aromatic ring),

104.15 and 96.83 (C-3, C-5).

Alternatively, the compound prepared in the previous example (6 g) was stirred for 16 hours and then heated at reflux for 4 hours and evaporated. The title product is obtained in the form of a sticky foam (6.2 grams) whose thin layer chromatography (Rf and eluents above) shows that the product is of sufficient purity for direct use in the next step.

In the same manner, the same product was prepared by solvolysis of the 2'-O-propionylester of the previous example.

Example 7 4 '-epi-9 - (^ leov3-)^Fc9 '] 4Tz [n4hu-'lio] ^ no (2rythromycin A (VI)

Method A

The product prepared in Example 2 (40 g) was dissolved in 600 ml of methanol. NaBHi (45 g) was added over 45 minutes while maintaining the temperature below 38 ° C. The reaction mixture was stirred for 64 hours and evaporated to give a thick slurry containing excess borohydride and the boronic ester complex with the product. The product thus obtained is partitioned between 500 ml of dichloromethane and water and the following sequence is repeated three times: the pH is adjusted to constant pH 2.5 with dilute hydrochloric acid, the mixture is stirred vigorously for 25 minutes and the aqueous phase is separated, soldered with 500 ml of fresh dichloromethane. The pH is adjusted to 9.5 with dilute NaOH and the dichloromethane phase is separated. The dichloromethane phase, pH 9.5, is mixed with 500 ml of fresh water and the sequence repeated. On the third repetition, the dichloromethane phase pH 9.5 was dried over sodium sulfate and evaporated in vacuo. The crude title product is obtained as a foam, 34 g, which is recrystallized from 150 ml of hot isopropyl ether, cooled, and diluted with 300 ml of pentane. This gives the purified title product, 25.8 g, as white crystals. TLC Rf = 0.5 (9: 1, chloroform: diethylamine) Rf = 0.1 (90: 10: 1, CHCl3:

: CH: OH: conc. NHiOH], mp 170-180 ° C,

1HNMR (CDCl3) δ:

2.26 (6H, s, (CH 3) 2 -),

3.29 (3H, s, cladinose CH3O-),

13CNMR (CDCl 3, / CH 3 / dSi internal standard) ppm:

179.44 (lactone C = O),

103.57 and 96.70 (C-3, C-5);

41.50 ((CH: R) -N).

Method B

The non-chromatographed title product of the previous example (6.2 g) was dissolved in 200 ml of ethanol and hydrogenated at 12.5 Raney-nickel at 40 psi for 18 hours. The reaction mixture is filtered, 20 g of fresh Raney nickel are added and hydrogenation is continued for a further 4 hours. Again, filtration and addition of fresh catalyst are performed and hydrogenation is continued for a further 16 hours. Filtration and evaporation of the filtrate gave the crude title product as a white foam. The crude product was partitioned between dichloromethane and saturated sodium bicarbonate and the organic phase separated, dried over sodium sulfate and evaporated. The title product was obtained as a second white foam (3.6 g) which was crystallized as described above to give the purified title product, 955 mg, with physical properties identical to the product prepared according to Method A.

Example 8

4 &amp; apos; -epi-9-deoxo-9a-hydroxy-9a-aza-9 &lt;a-hornoerythromycin A

3'-N-oxide (X)

While stirring at atm. In the nitrogen spheres, the product of the previous example (3.0 g) was dissolved in 15 ml of a 1: 1 mixture of tetrahydrofuran and methanol. 30% Hydrogen Peroxide (5 ml) was added. After an additional 30 minutes, the reaction mixture was carefully poured into CH 2 Cl 2 and water 1: 1 containing Na 2 SO 4 (exothermic) The pH of the reaction mixture was 9. The aqueous phase was washed with fresh dichloromethane and then with ethyl acetate. The organic phases were combined, dried over sodium sulfate and evaporated to give the title product. Yield 2.7 g. TLC Rf 0.15 (60: 10: 1, CH 2 Cl 2: CH 3 OH: conc. NH 4 OH);

1HNMR (CDCl3) &lt;5:

3.21 (6H, s, / CH 5/2 N, O),

3.38 (3H, s, cladinose CH3O-),

MS: major peaks at m / e 576 (ion from desosamine fragmentation at C-5), 418 (N-hydroxyaglycone ion minus both sugars). Both. the peaks are diagnostic for the part of the N-OH molecule with aglycone.

Example 9

4^at-epi-9-deoxo-9α-methyl-9α-aza-9α-homoerythrcmycin A

3'-N-oxide (XI)

The product of the previous example (2.6 g, 0.0034 mol) was dissolved in 100 ml of dichloromethane. Under vigorous stirring, K 2 CO 3 (37.5 g, 0.271 mol) was added followed by methylpodide (19.3 g, 8.5 mL, 0.136 mol) and the reaction mixture was stirred for 20 hours. Yield 2.9 g, thin layer chromatography Rf 0.3 (60: 10: 1, CH 2 Cl 2: CH 3 OH:: conc. NH 4 OH), R f 0.15 (90: 10: 1, CH 2 Cl 2: CH 2 OH: conc.

The title product, prepared

219 in this way - (2.8 g) was further purified by chromatography on 85 g silica gel - using - 90: 10: 1 CH 2 Cl 2: CH 2 OH: conc. NH 4 OH as the eluent. This removes minor - more polar impurities. Isolate - 0,87 g of product,

@ 1 H NMR (CDCl3) .delta .:

2.32 (3H, s, aglycone CH3-N-),

3.20 (6H, s, (CH3) N-O),

3.37- (3H, s, Cladinose CH3O-J).

Example 10

4'-epi-9-deoxo-9α-methyl-9α-aza-9α-homoerythromycin A (IV)

Method A

The product of Example 7 (0.706 g, 0.96 mol) was dissolved in 20 ml of chloroform. Formaldehyde (37%, 0.078 mol) was added - and then - formic acid - (0.03 ml) and the reaction mixture was stirred for 4 hours and then heated at reflux for 7 hours. The reaction mixture was cooled, 30 ml of water was added, and the pH was adjusted to -9 with 6N NaOH. The organic phase was separated, dried over sodium sulfate and evaporated in vacuo to give the title product as a white foam. Yield 0.7 g of product which, after crystallization from hot ethanol-water, yields 302 mg of product mp 153 ° C and recrystallization from hot ethanol-water mixture gives 246 mg of product mp 155 ° C. TLC Rf = -0.55 (60: 10: 1, CH 2 Cl 2: CH 2 OH: conc. NH 4 OH), Rf 0.6 (9: 1 - CHCl 5: diethylamine);

4HNMR (CDCl3) δ:

2.29 (9H, broad s, aglycone N-CH3 and desosamine (/ CH3 / 2N-),

3.31 (3H, s, cladinose CH3O-);

13CNMR (CDCI3, / СИз / tSi internal standard) ppm:

178,89- (lactone C = -O),

102.63 and 95.15 (C-3, C-5 ·),

40.38 - [(CH 3) 2 N -);

MS: major peaks at m / e 590 (M-methylaglycone-desosamine ion by Cladinase cleavage - to Cl '), 416 [N-methyl aglycone ion (minus -oba - sugars by Cl' cleavage, - C-5)], 158 ) major peak, a fragment derived from cd desosamine j.

Method B

Non-chromatographed product (0.242 g) and 10% palladium on carbon. (0.4 g) is mixed in 15 ml of 95% ethanol and the mixture is hydrogenated for 1 hour at 50 psi. The catalyst was filtered off and the filtrate was evaporated. The title compound was obtained as a white foam, 160 mg, which after crystallization from ether / pentane, 124 mg, and recrystallization from ethanol / water, 95 mg, gave a product with physical properties identical to the product prepared by Method A.

Method C

The title product, chromatographically purified, of the previous example (319 mg), and Rahey-Nickel (1.5 g, -50% wet) are mixed in 20 ml of ethanol and hydrogenated under a pressure of 0. , 34 MPa for 1.5 hours. The catalyst is filtered off and the mother liquors - evaporated - to dryness. - 205 mg of the title product is obtained which, in its physical properties, is identical to that of the product prepared by method A.

Example - - - 11,

2'-O-acetyl-9-deoxo-9α-methyl-9α-aza-9α-homoerythromycin A

The product of Preparation 5 (2.5 g 3.34 mmol) was stirred with acetic anhydride (0.339 mL, 3.60 mmol) in 30 mL of dichloromethane for ~ 4 hours. The reaction mixture is evaporated under vacuum and the residue is dissolved in 50 ml of ethyl acetate, treated with 50 ml of water and the pH is adjusted to 9.5 with 1N NaOH. The aqueous phase was separated and washed with 20 ml of fresh ethyl acetate. The organic phases were combined, dried over sodium sulfate, evaporated and dissolved in 30 ml of chloroform and evaporated again. The title product is obtained as a dry solid. Yield 2.82 g.

AINMR (CDCl3) δ: includes

3.31 (C4 "-OCH 3),

2.28 (N-CH 3),

2.25 (N- / CH 3/2) a

2.0 (2'-OCOCH 3).

Example -1 ad 12

2'-O-acetyl-4'-deoxy-4'-oxo-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin A (VIIIa)

The title product of the previous example (2.5 g, 3.2 mmol) and dimethyl sulfoxide (0.38 mL, 5.23 mmol) were dissolved in 90 mL of dichloromethane and cooled to -70 degrees Celsius. Trifluoroacetic anhydride (0.72 mL, ,95 4.95 mmol) was added via syringe to keep the temperature below -50 ° C and the mixture was stirred at -60 ° C for 50 min. Triethylamine (1.54 mL, 11 mmol) was added

4 1 β 6 9 syringe and the temperature is maintained below -50 ° C during the addition. The mixture was then warmed to 0 ° C, diluted with water and the pH adjusted to 9.5 by addition of dilute sodium hydroxide. The organic phase was separated, dried over sodium sulfate and foamed to give the title product [2.5 g). The foam is chromatographed on silica gel with 10: 1 chloroform: methanol and 3 fractions are collected by thin layer chromatography analysis. The pure product from fraction 1, 1.7 g, was dissolved in chloroform, diluted with water, adjusted to pH 4 with dilute hydrochloric acid and the aqueous phase separated, diluted with fresh chloroform, adjusted to pH 8 with dilute sodium hydroxide and the organic phase separated . The last aqueous phase was extracted with three portions of fresh chloroform. The last four phases were combined, washed again with water, dried over sodium sulfate and evaporated in vacuo to give the title product. Yield 0.98 g of thin layer chromatography Rf-0.7 (5: 1: 0.1, CHCl 3: CH 3 OH: NH 4 OH),

1 HNMR (CDCl 3) δ (ppm):

2.05 (s, 3H, COCH 3),

2.26 (s, 6H, N / CH 3/2),

2.33 (d, 3H, NCH 3) a

3.33 (d, 3H, OCH 3).

Example 13

4'-deoxy-4'-oxo-9-deoxo-9α-methyl-9α-aza-9α-homoerythromycin A (Vila)

The title compound (0.93 g) was dissolved in methanol. After 20 minutes the mixture was evaporated to give the title product. Yield 0.74 g, MS 746.4, 588.4, 573.4, 413.3, 158.1, 125.1,

1HNMR (CDCl3) &lt;5 (ppm):

5.5 (t, 1H, Cl-H),

4.6 (q, 1H, C5-H),

3.35 (s, 3H, OCH 3),

2.38 (s, 3H, NCH 3),

2.30 (s, 6H, N (CH3) 2).

Example 14

4^at-epi-9-deoxo-9α-methyl-9α-aza-9α-homoerythromycin A (1V)

The title product of the preceding example (0.25 g) and 250 mg of Raney-nickel were combined in 20 ml of ethanol and hydrogenated at 20 psi for 4 hours. The catalyst was filtered off and evaporated in vacuo to give an oil which crystallized on standing. The title product was isolated by trituration in isopropyl ether and filtration. Yield 0.13 g of product having the same characteristics as the product of Example 10.

Preparation 1

4'-epi-erythromycin A

A suspension of 100 g of Raney-nickel in 1 liter of absolute ethanol containing 100 g of 4 &amp; apos; -deoxy-4 &amp; apos; -oxoerythromycin A (U.S. Pat. No. 4,510,220) was shaken under a hydrogen atmosphere overnight at room temperature and pressure. . The spent catalyst is filtered through diatomaceous earth and the filtrate is concentrated to 300 ml in vacuo. Water (700 ml) and the concentrated filtrate were added and the resulting milky solution was heated on a steam bath. A small amount of ethanol is added to prevent gum formation during precipitation from the hot solution. After stirring at room temperature for 2 hours, the product was filtered off and dried. 57.6 g of product are obtained, and the filtrate is concentrated in vacuo to turbidity. The mixture was then stirred for one hour, then filtered and dried. Yield 21.4 g.

The crystalline fractions obtained are combined and have a melting point of 141-144 ° C.

Their 1 H NMR spectrum (CDCl 3) shows absorption at:

3.3 (3H, s)

2.3 (6H, s);

1.4 (3H, s) ppm.

Preparation 2

Oxime erythromycin hydrochloride

Under nitrogen, erythromycin A (500 g, 0.681 mol) was dissolved in pyridine (2.787 kg, 2.850 L, 35.29 mol). Hydroxylamine hydrochloride (1.183 kg, 17.02 mol) was added and the mixture was stirred for 22 hours, then evaporated to a thick slurry which was filtered while rinsing with isopropanol. The combined filtrates were again evaporated to a thick waxy mass which crystallized by trituration with 2 L of water. The product 615 g (slightly wet, used in the next step without drying). TLC Rf 0.45 (60: 10: 1, CH 2 Cl 2: CH 3 OH: conc. NH 4 OH).

Using the same procedure, 5 g of erythromycin A is converted to the dry title product. Yield 4.5 g at least 95% pure by 13C NMR. Recrystallization of 1 g of 10 ml of methanol and 30 ml of isopropyl ether gives 725 mg of mp 187 ° (dec.), (Literature mp 188-191 ° C, Massey et al., Tetrahedron Letters, pp. 157-160, 1970),¹³CNMR (DMSO-d6, / CH3 / 4S1 internal standard) ppm, 174.35 (lactone C = O), 168.78 (C = N -), 101.0 and 95.46 [C-3, C-5] ).

Preparation 3

9α-aza-9α-homoerythromycin A

Following the procedure of Example 2, the slightly wet title product of Preparation 2 (615 g, calculated on 506 g, 0.613 mol of dry matter) was converted to the crystalline title product to evolve the gas observed upon addition of hydrogen 241069 sodium carbonate. Yield 416 g,¹³CNMR (CDCl 3, CDCl 3 internal standard) ppm

177.54 (lactone C-O), 163.76 (amidic C = O), 102.28 and 94.20 (C-3, C-5), 40.13 [(CH3) zN-J].

Preparation 4

9-deoxo-9-aza-9α-homoerythroinycin A

By reduction of NaBH4 by the method of Kobrehe-