Process for preparing epimeril aza homoerytheomycin a derivati-ves and intermediates therefor

DESCRIPTIVE MEMORY

This invention relates to antibacterial 4-epi-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin A, their pharmaceutically acceptable salts, and intermediates useful in their preparation from erythromycin A.

Erythromycin A is a well-known macrolytic antibiotic having formula (I) which finds wide clinical use.

PFIZER INC.

PROCESS FOR THE PREPARATION OF AZA EPIMERIC DERIVATIVES_ —HOMOERYTHROMYCIN A AND ITS INTERMEDIATE PRODUCTS

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This therapeutic compound is the 4-epimer derived from erythromycin A above of formula (II), the

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-3 (II) B = methyl (III) E = hydrogen subject to Belgian patent 892,557, as well as my co-pending US serial no. 589,401, filed July 19, 1982. In this Belgian Patent, the compound of formula (II) is termed as the N-methyl derivative of 11-aza-10-deoxo-10-dihydroerotromycin A, a name subsequently coined by Kobrehel et al., US Patent 4,528,534 for the precursor compound of formula (III). For the aaa (nitrogen-substituted carbon) derivative of erythromycin A with the last enlarged (homo) ring, we prefer the name 9-oxos 9 -a-aza-9a-homoerythromycin A. This compound may also be referred to as 10-aza- 14-hexadecanoyl acid.

Some of these novel intermediates are also 4-epimers of previously known compounds. Thus 4-epi-9-deoxo-9a-aza-9a-homoerythromycin A is the 4-epimer of the above-mentioned compound of formula (III); and 4-epierythromycin Oxime is the 4-epimer of erythromycin A oxime of Djokic et al., US Patent 5-478,014.

4-Epierythromycin A is the subject of co-pending US patent application, U2 Series. 553-547, filed March 1, 1982 by Sciavolino et al.

This invention includes the 4-epi-9-deoxo-9a-methyl-9a-aza-r9a-homoerythromycin A antibacterial compound having formula (IV), its pharmaceutically acceptable salts, its pharmaceutical compositions, and a method of use thereof. treatment of bacterial infections in mammals.

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(IV) R = methyl, Z = Z = hydrogen (V) E = hydrogen, Z and Z¹ together = oxygen (VI) R = Z = Z¹= hydrogen '

This therapeutic compound (IV) discloses a relatively broad spectrum of antibacterial activity including erythromycin A susceptible organisms and further includes the most important respiratory pathogen Hemophillus influenzae. Its high oral absorption and its extraordinarily long half-life in vivo makes compound (IV) especially valuable in the oral treatment of bacterial infections in mammals.

*

This invention also includes intermediates useful in the art. Synthesis of 4-epi-9-d.esoxo-9a-methyl-9a-aza-9a-homoerythromioin A (IV) as follows (a) A compound selected from the group consisting of 4 '^,-epi-9a-^The-za - 9^The-Uomoerythromycin A and its 9-deoxo derivative with the above formulas (V) and (VI) respectively

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(b) 4-Epierythromycin A oxime.

(c) A compound selected from the group consisting of 9^The- Benzyloxycarbonyl-9-deoxo-4''-deoxy-4-oxo-4 ”-9a-aza-homoerythromycin A of formula (VII); 9-deoxo-4-deoxy-4-oxo-9a-methyl-9a-aza-9a-homoerythromycin A of the formula (Vila); and the corresponding 2'-O- (C 2 -C 6) alkanoyl derivatives of the formulas (VIII) and (VIIIa). Acetyl is the preferred value of 2'-O- (C6 -C6) alkanoyl.

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ca och₃

P (VII) R 'benzyloxycarbonyl, R = H (VIII) R3 benzyloxycarbonyl, R²= (C2 "Cj) alkanoyl (Vila) R¹= methyl, R²= H (VIIIa) R 2 = methyl, R 4 (C 1 -C 6 -C 6) alkanoyl (d) A compound selected from the group consisting of 2'-O-acetyl- and 2'-O-propionyl-9-deoxo-9a benzyloxycarbonyl-9a-aza-9a-homoerythromycin A of formula (IX). The 2'-0-aeetil derivative is of particular value.

-ζ&gt; -

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(IX) R 4 Uenzyloxycarbonyl, R² a (C 6 -C 6 alkanoyl and (e) A compound selected from the group consisting of 4-deoxo-9a-methyl-9a-aza-9a-hoapherythromycin A 5'-N-oxide of formulas (X) and (XI), respectively.

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(X) R4 = hydroxy (XI) R4 = methyl

The antibacterial compound of this invention, 4-epi-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin A (IV), is readily prepared by a number of routes from erythromycin A. which are variously derived via novel compounds known as intermediate, include intrinsic transformations as follows:

(A) 0-4 epimerization;

(B) ring widening with introduction of 9a-nitrogen;

(C) removal of the 9-oxo group; and (D) 9a-N-methylation;

together with any necessary or fecultative introduction or removal of protecting groups. Not preferred are either of the following transformation sequences: (A) (B) (C) (D), (B) (A) (C) (D) or (B) (C) (D) (A). The various intermediates and the final product are isolated by standard handling methods (for example), extraction, precipitation, evaporation, chromatography, crystallization.

(A) (B) (0) (D)

Operational sequence (A) (B) (0) (D) includes the initial conversion of erythromycin A (I) to 4-epierithromycin A according to the method of Sciavolino et al. (supra). The latter is then converted, in virtually quantitative production, to 4-epierythromycin A

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oxime by reaction with hydroxylamine or preferably a hydroxylamine salt such as hydrochloride. Under preferred conditions, recent discoveries, at least one molar equivalent is used, usually an excess, e.g., equivalent, of hydroxylamine; in an excess of weakly basic tertiary amine (preferably pyridine) as solvent; at a temperature ranging from 0-50 °, conveniently at room temperature.

The resulting 4-epierythromycin oxime is rearranged into the 4-epi-9a-aza-9a-homo derivative (V) via a Beckman rearrangement. Preferred conditions utilize an excess (e.g. 5-4 molar equivalents) of an organic sulfonyl chloride, preferably methane sulfonyl chloride, which is reacted with the oxime (as a free base or as an acid salt). ) in a mixture of a lower ketone (e.g. methyl ethyl ketone) and water containing a large molar excess of sodium bicarbonate at a temperature of 0-50 ° C, preferably 0-50 ° C.

The C-9 carbonyl amide of (V) is then conveniently reduced to the corresponding dihydro derivative i.e. 4-epi-9-cesoxo-9a-aza-9a-homoerythromycin A (VI) by reduction with sodium borohydride (preferably in excess to force the reaction to complete in a reasonable period of time, but at least equivalent). The reduction is carried out in a suitable protic solvent, such as a lower alkanol (preferably methanol) at 0-50 ° (preferably at or below 58 °). 0 Excess NaBH4 is carefully decomposed by quenching the reaction into dilute aqueous acid.

Final methylation to afford compound (IV) is by reduction 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

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The reaction is preferably carried out with at least one equivalent of each of the formaldehyde and formic acid in an inert reaction solvent at 20-100 ° C. The preferred solvent is chloroform. In this solvent, the reactants are conveniently combined at room temperature and then heated under reflux to force the reaction to complete.

Alternatively methylation of (VI) to (IV) is effected by oxidatively protecting the dimethyl amino group as its N-oxide (while forming the 9a-N-hydroxy derivative), methylating with methyl iodide with (at least in part) 9a-N-simultaneous deoxygenation, and reduction of the resulting 9a-methyl-5-N-oxide. The oxidation of (VI) is effected rapidly by reaction with hydrogen peroxide, usually in excess of the two required minimum molar equivalents, in an inert reaction solvent 10 50øC, conveniently at room temperature. In this way 9α-hydroxy-3'-β-oxide (X) is formed. The latter is methylated and deoxygenated in (XI) with methyl iodide conveniently in an inert reaction solvent (e.g. methylene chloride) at 0-5 ° C. (conveniently at room temperature), preferably in the presence of an insoluble base solvent which will neutralize the acid formed (e.g. HI when methyl iodide is the methylating agent). With methylene chloride as a solvent, an excess of potassium carbonate is the basis of choice. Thus the excess base and the formed sodium iodide are completely removed by simple filtration before isolation of a 9α-methyl-3'-α-oxide (XI). Finally, removal of the 3'-N-oxide group is effected rapidly by hydrogenation over a Ranay noble or nickel metal catalyst. In this hydrogenation, temperature and pressure are not of critical importance, for example, suitably 0-100 ° C and a pressure ranging from subatmospheric to 100 atmospheres or more. More convenient are ambient temperature and moderate pressures, for example 2-8 atmospheres. Suitable noble metal catalysts include palladium, rhodium and platinum, supported or unsupported type, well known in the art.

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catalytic hydrogenation technique. Preferred catalysts are palladium on a carbon and nickel

Raney

(Β) (A) (0) (D)

Operational sequence (Β) (A) (C) (D) includes the initial conversion of erythromycin A (I) to 9-deoxo-9a-aza-9a-homoerythromycin (III) by erythromycin A oxime and 9α aza-9a-homoerythromycin according to the method of Kobrehel et al. (supra). In this context, the novel process described above for 4-epierythromycin A oxime is advantageously used for the preparation of the intermediate erythromycin A oxime.

The 2'-hydroxy group of compound (III) is first protected as its acetate or propionate ester. Acylation is selectively performed by reacting compound (III) with a limited excess of acetic or propionic anhydride in an inert reaction solvent (e.g. methylene chloride) at Q ~ 20 ° C. (conveniently at room temperature). 0 Limited excess anhydride is used to compensate for reagent consumed in secondary reactions, for example undesired acylation of other groups, particularly 9a-azoxo.

The resultant 2 0 (O ^ -0303) alkanoyl derivative is then protected on nitrogen with a benzyloxycarbonyl group. Thus compound (IX) is formed by reacting the above 2'-ester with carbobenzoxy chloride in an inert reaction solvent in the presence of a base. Particularly well-suited are the Schotten-Baumann conditions, that is, the reaction of the 2'-ester with acid chloride under aqueous, alkaline conditions eg aqueous tetrahydrofuran, maintaining pH 7.5-8.5 with dilute NaOH while acid chloride is added and while the reaction is continued. The temperature is not of critical importance, but will generally range from 0-50 ° C, conveniently at room temperature.

Compound (IX) C-4 hydroxyl is then oxidized to C-4-oxo (VIII) by the action of oxalyl chloride / dimethyl sulfoxide at a low temperature (-40 to -80 ° C) in an inert reaction solvent. (e.g. methylene chloride) followed by treating the cold reaction mixture with an excess of tertiary amine (e.g. triethylamine). 0 Ester alkanoate protecting group is removed by solvolysis, preferably by contact with excess methanol at 0-100 ° C thus forming compound (VII).

Hydrogenation over Raney nickel catalyst using the conditions described above converts compound (VII) to 4-epi-9-deoxo-9a-aza-9a-homoerythromycin A (VI). The latter is converted to a 9a-N-methyl (IV) derivative according to one of the alternative methods as described above.

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

This operational sequence includes the initial conversion of erythromycin A to the above-mentioned compound of formula (II) according to my aforementioned co-pen application, using methods detailed in the following section of the Preparation. The C-4 epimerization is then performed according to the steps and methods described above. 0 2'-hydroxy group is protected by acylation, the 4-hydroxy group being oxidized to 4-oxo group, preferably replacing oxalyl chloride with trifluoroacetic anhydride; the acyl protecting group is removed; and the 4-oxo group is catalytically hydrogenated to give the desired 4'-epimeric hydroxy group. In this case, the preferred catalyst is Raney nickel.

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Since the compound (IV) of this invention contains two basic nitrogen atoms, pharmaceutically fluctuating mono- and diacid addition salts are formed by contacting the free base (IV), respectively with substantially one equivalent of the acid or at least two. acid equivalents. Salts are generally formed by combining the reactants in an inert reaction solvent; If the salt does not precipitate directly, it will be isolated by concentration and / or addition of a non-solvent. Suitable pharmaceutically acceptable acid addition salts include, but are not limited to, those formed with HCl, HBr, ΗΗΟΗΗΟ, HgSO ^, HOgCCHgCHg C00.₂H, cis- and trans-H0₂CCHCHC0₂H, CH 2 SO 4 H and ja-CH 2 C 6 H 4 SO 4 H.

The antibacterial activity of the compound of formula (IV) is demonstrated by measuring its minimum inhibitory concentration (MIC) in mcg./ml. against a number of microorganisms in heart and brain infusion broth (IOC). Twelve double dilutions of the test compound are generally used, with the initial concentration of the test product ranging from 50 to 200 mog / ml. Susceptibility (CIW) of the test organism is accepted as the lowest concentration capable of producing complete inhibition of growth as assessed by the naked eye. Comparison of h activity^no-epi-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin A (IV) with that of control erythromycin A is indicated in duplicate in Table I.

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TABLE I

In Vitro Aetivity of Compound (IV)

Duplicate CIM Values

Day 1 day 2

<td></td><td colspan="2">THE</td><td>B</td><td>THE</td><td>B</td>

<td>Staph aur.</td><td> 005</td><td> 0.05</td><td> 0.20</td><td> 0.05</td><td> 0.39</td>

<td></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>Staph epi</td><td> 111</td><td> 0.05</td><td> 0.10</td><td> 0.05</td><td> 0.20</td>

<td>Strep. do</td><td> 006</td><td> 0.78</td><td> 1.56</td><td> 0.78</td><td> 0.78</td>

<td>Strep. pyog.</td><td> 203</td><td> 0.025</td><td> 0.025</td><td> 0.025</td><td> 0.025</td>

<td>Strep. 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>(The)</td><td> 6.25 '</td><td>(The)</td><td> 6.25</td>

<td></td><td> 129</td><td>(The)</td><td> 1.56</td><td>(The)</td><td> 6.25</td>

<td></td><td> 266</td><td>(The)</td><td> 3.12</td><td>(The)</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>Kleb. pn.</td><td> 009</td><td>(The)</td><td> 12.5</td><td>(The)</td><td> 12.5</td>

<td></td><td> 031</td><td>(The)</td><td> 12.5</td><td>(The)</td><td> 12.5</td>

<td>Kleb. oxy.</td><td> 024</td><td>(The)</td><td> 12.5</td><td>(The)</td><td> 12.5</td>

<td>Past very many</td><td> 001</td><td> 1.56</td><td> 0.10</td><td> 1.56</td><td>0.Ϊ0</td>

<td>Serr. sea.</td><td> 017</td><td>(The)</td><td> 50</td><td>(The)</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>(The)</td><td> 12.5</td><td>(The)</td><td> .12.5</td>

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

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

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TABLE I (Pont.)

In Vitro Aetivity of Compound (IV)

L ·, ·

H. influ.

Duplicate CIM Values

<td></td><td>Day 1</td><td></td><td>Day</td><td> 2</td>

<td></td><td>THE</td><td>B</td><td>THE</td><td>B</td>

<td> 042</td><td> 1.56</td><td> 0.39</td><td> 1.56</td><td> 0.39</td>

<td> 051</td><td> 3.12</td><td> 0.39</td><td> 3.12 ·</td><td> 0.78</td>

<td> 073</td><td> 3.12*</td><td> 0.39</td><td> 3.12</td><td> 0.78</td>

<td> 078</td><td> 1.56</td><td> 0.39</td><td> 1.56</td><td> 0.39</td>

<td> 081</td><td> 3.12</td><td> 0.39</td><td> 3.12</td><td> 0.78</td>

(a) larger than 50.

Erythromycin A as a control

B Compound (IV)

Additionally, compound (IV) is tested in vivo by the well-known mouse protection test or by a microbiological (bioassay) determination of serum levels in a number of mammals (eg, rat, rat, dog). ) When rats were used as the test species, compound (IV) was found to be exceptionally well absorbed after oral dosing, providing exceptionally high and long-lasting serum levels.

The:

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For the treatment of systemic infections in animals, including man, caused by susceptible microorganisms, compound (IV) is dosed at a level of 2.5-100 mg./kg. per day, preferably 5.50 mg./kg./day, in divided doses or preferably in a single daily dose. The variation in dosage will be made depending on the individual and the susceptibility of the microorganism. These compounds are orally or parenterally dosed, the preferred route being oral. The susceptibility of microorganisms. isolated in clinics is routinely tested in clinical laboratories by the well-known plate-disc method.

Compound (IV) is generally the compound of choice when it relates to a relatively large zone of inhibition against the bacterium causing the infection to be treated.

Preparation of optimal dosage forms will be by methods well known in the pharmaceutical art. For oral administration, the compounds are formulated alone or in combination with pharmaceutical carriers such as inert solid diluents, aqueous solutions or various non-toxic solvents in dosage forms such as gelatin capsules, tablets, powders, tablets, syrups, etc. . Such vehicles include water, ethanol, benzyl alcohol; glycerin, propylene glycol, vegetable oils, lactose, starch, talc, gelatins, gums and other well known carriers. The parenteral dosage forms required for the aforementioned systemic use are dissolved or suspended in a pharmaceutically acceptable carrier such as water, saline, sesame oil, etc. Agents that enhance the qualities of suspension and dispersion may also be added.

For the topical treatment of superficial infections in animals, including man, caused by susceptible microorganisms, compound (IV) is formulated by methods well known in the pharmaceutical art to lotions, ointments, creams, ointments, gels, or others in varying concentrations. 5-200 mg./cmj of the dosage form, preferably ranging from 10-100 mg./cmj. The dosage form is applied at the site of ad libitum infection, usually at least once a day. This invention is illustrated by the following examples. However, it should be well understood that the invention is not limited by the specific details of these examples. Unless otherwise specified, all operations were performed at room temperature; all solvent removal was performed in vacuo from a bath at 40 ° or less; all temperatures indicated are in degrees Centigrade; all thin layer chromatography (TLC) was performed on commercially available silica gel plates (using the eluent indicated in parentheses); and all solvent ratios are by volume. THE¹ It is used for tetrahydrofuran, and DMSO is used for dimethyl sulfoxide.

EXAMPLE 1

4-Epierythromycin A Oxime / 4-Epimer cLe Oxime (1) 7

4-Epierythromycin A (50 g.,

0.0646 mole) was dissolved in 265 ml of pyridine. Hydroxylamine hydrochloride (112.2 g, 1.615 mole) was added and the slurry stirred for 16 hours. The reaction mixture was purified to a thick slurry, diluted with 500 ml isopropanol, shaken well and filtered with 3 g. x 100 ml. of isopropanol for washing. 0 The filtrate and the washings were combined, purified to a water-soluble foam, and triturated with ether to afford the crude title product as a hydrochloride salt (100 g). The latter was purified by distributing it between CH 2 Cl 2 and aqueous NaHCO 3 adjusted to pH 9.5 with dilute NaOH.

The aqueous layer was separated and washed with ethyl acetate and then ether. All organic layers were combined dried (NagSO4) and purified to afford the title product as a white foam, 59.5 g.

ccf Rf 0.5 (CH₂C1₂: CH₂OH: RH₄OH conc. 60: 10: 1); 1 H NMR (CDCl3) delta 2.31 / 6H, s, 3.32 (3Ξ, s, cladinosis CH2 O-).

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EXAMPLE 2

4^ll-Epi-9a-aza-9a-homoerythromycin A (V) title product from the previous Example (59.2 g, 0.0787 mol) was dissolved in 400 ml of acetone. A RaHCO3 slurry (60 g) in 225 ml HgO was added. Methanesulfonyl chloride (36) 3 g, 24.5 ml) in 50 ml of acetone was added portionwise over 10 minutes, while maintaining the temperature below 30 ° by means of a cooling bath. The mixture was stirred for 4.5 hours, removing acetone and CH 2 Cl 2 (400 mL) was added to the aqueous residue, and the pH was adjusted to 5) 6 with 6N HCl. The aqueous layer was separated, washed with two additional portions of CB 2 Cl 2 θ then adjusted to pH 9.5 with 6N NaOH. The basic solution was extracted 2 x with fresh CBCl 3, 1 x with ethyl acetate and 1 x with ether. The basic organic extracts were combined, dried (N 2 SO 4) and purified to afford the title product as a solid. of an ivory foam, 41 g, ccf Rf 0.4 (CH₂C1₂: CH₅OH: RH₄0H conc. 60: 10: 1); ^ Ήποη (CDCl 3) delta 2.27 / ~ 6H, s, (0Η₅)₂Ν₌7.29 (3H, s, cladinose CH 2 O-);

Crmn /^-CDCl3, (CH3)₄ internal standard Si7 ppm 177.24 (lactone 0 = 0) 163.53 (amide 0 = 0), 102.29 and 95.24 (C-3, C-5), 40.22 / “(ch₃)₂no_z7.

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EXAMPLE 3

(111) 7 2'-β-Acetyl-9-deoxo-9a-aza-9a-homoerythromycin A / ”2'-0-Acetate

9-Deoxo-9a-aza-9a-homoerythromycin Î ± (10 g, 0.0156 mol; (lil); US Patent 4,528,554) was dissolved in 150 ml of CH2 Cl2. Acetic anhydride (1.59) was added. S, 1.28 ml 0.0156 mol) and the mixture was stirred for 5 hours. Acetylation was monitored by tlc; To force the reaction to completion, 0.25 ml of acetic anhydride and then 0.5 ml of acetic anhydride were added, with further stirring respectively.

1.5 and 1 hours. The reaction mixture was diluted with H₂0 and pH was adjusted to 11 with dilute NaOH. The organic layer was separated, dried (NaSO4), and purified to a foam, 11.5 S ·^THE foam (10 g) was chromatographed on 500 g of silica gel with CH₂C1₂: CH 3 OH 9: 1 as eluent and tlc monitoring. A less polar impurity (5.6 g) was eluted, followed by the purified title product, isolated as a white foam, 2 g TLC Rf 0.2 (CH3 Cl2 CH3 OH: NH4 OH conc. 90: 10: 1) • nmr (CDCl3) delta 2.02 (5H, s, 0-2 'θ it

-OO-CH ^)

2.26 / 6H, s, (CH₅)₂N₌7, 5.35 (3Ξ, s, cladinosis 40-).

By substituting propionic anhydride with acetic anhydride, the corresponding derivative 2 was prepared by the same method.¹-O-propionyl.

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Example 4 * -O-Acetyl-9-deoxo-9a-benzyloxycarbonyl-9a-aza-9a-homoerythromycin A / “(IX), R = acetyl7 title product from the previous Example (1.7 g, 0.00219 mole) It was dissolved in 70 ml of 5: 2. The pH was adjusted to 8 with diluted NaOH.

carbobenzoxy chloride (0.51 g, 0.427 mL, 0.003 mol) was added and the mixture was stirred for 2 hours with further diluted NaOH added as needed to maintain pH 8. As tlc indicated incomplete reaction, added more carbobenzoxy chloride (0.3 ml), and the reaction was continued for 3 hours while still maintaining pH 8. The reaction was quenched with co. NaHCO 3 and ethyl acetate, the pH was adjusted to 9.6 and the aqueous layer was washed with CH2 Cl2, the organic layers were combined, dried (NagSO4) and purified to give a foam, 2.4 g. The foam was chromatographed on 85 g of silica gel, eluting with conc. 170: 10: 1. The processed pure fractions were combined to a foam, mixed with CH 2 Cl 2 and concentrated to the crystallized title product, 1.2 g, mp 122 °; tlc Rf 0.4 (CH3 Cl2: 2.27 / 6H, s, (OH₅)₂N_;;7.35 (3H, cladinosis CH 2 O-); ¹⁵Internal Standard Si / ppm 176.31 (lactone 0 = 0), 169.36 (0-2 'ester 0 = 0), 157.10 (carbamate 0 = 0); 137.0, 127.55 and 127.92 (aromatic ring); 40.6 / “(CH3 N;

By the same method, 0 derivative

The 2'-O-propionyl of the previous Example is converted to the corresponding 2'-O-propionyl-9α-benzyloxycarbonyl derivative.

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EXAMPLE 5'-O-Acetyl-9a-benzyloxycarbonyl-9-deoxo-4p

-oxo-9a-aza-9a-homoerythromycin A (fVII), R = acetyl?

Oxalyl chloride (4.57 g - 3 ml - 0.0544 mole) was dissolved in 25 ml of CH 2 Cl 2 and cooled to -60 °. DMSO (6.70 g, 6.09 ml, 0.0856 mole) in 9 ml CHgClg was added. After keeping the mixture at -60 ° for 10 minutes, the title product of the previous Example (5-2 g, 0.00572 mol) in 16 ml of

After a further 25 minutes at -60 °, triethylamine (17.3 g, 25.9 ml, 0.172 mol) was added and the mixture was warmed to room temperature, diluted with 5θ ml HgO and excess RaUCO3. . The organic layer was separated, dried (Na2 SO4) and purified to afford the title product as a gummy foam, 6.8 g; ccf Rf 0.6 (0Ξ20ΐ2: 0Ξ20Ξ: ΟΞ ^ ΟΞ conc. 90: 10: 1)¹HRMN (CDC1₅) delta 2.05 (5H, s 0-2 '

II

-OC-OH ^)

2.25 / 5.3, (CH₅)₂N, 7.52 (5H, s, cladinosis CH2 O-), 7.57 (5, s, aromatic protons); MS: higher peaks at m / e 536 and 518 / N-beuzyloxycarbonyl aglycone ion (minus both sugars via cleavage at 0-1, 0-5) 7? 200 (base peak, neutral sugar-derived fragment). This intermediate is preferably used immediately in the next step).

Similarly, the corresponding derivative 2^,-0-propionyl-4-oxo is prepared from the 2'-O-propionyl compound of the previous Example.

act;

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-21EXAMPLE 6

9a-Benzyloxycarbonyl-9-de-soxo-4-de soxy-4-oxo-9a-aza-9a-homoerythromycin A (VII)

The title product of the previous Example 1.0 g was stirred in 25 ml of methanol for 65 hours, and then purified to a foam. The foam was mis. NaHCO 3 washed with saturated NaHCO 3, and purified again to give a second foam. The second foam was chromatographed on 20 g of silica gel using 13: 1 CH2 Cl2 ECH3 OH as eluent. The pure product fractions were combined and processed to afford the purified title product as a foam, 336 mg; tlc Rf 0.4 (03201?:: CH? OH: 1?₄0H conc. 90: 10: 1);¹⁵Crmn / CDCl 3, (internal standard CH 2 Si 7 ppm 210.87 (0-40 = 0), 176.03 (lactone 0 = 0), 157.41 (carbamate 0 = 0); 136.31, 128, 2 and 128.0 (aromatic ring);

104.15 and 96.83 (0-3.0-5).

Alternatively, the title product of the previous Example (6 g) was stirred for 16 hours, then refluxed for 4 hours and processed to give the product as a gummy foam, 6.2 g, layer chromatography. (Rf and eluent as mentioned above) indicated sufficient purity to be used directly in the next step.

Similarly, the same title product is prepared by solvolysis of the 2'-O-propionyl ester of the previous Example.

EXAMPLE 7

<img file="PT77645B_D0025.tif" />

4^ll-Epi-9-deoxo-9a-aza-9a-homoerythromycin A (VI)

Method The title product of Example 2 (40 g) was dissolved in 600 ml CH 2 OH. NaBH ^^ S? g) for 45 minutes keeping the temperature below 58 °. The reaction mixture was stirred for 64 hours, then purified to a paste containing excess borohydride and boro ester complex of the product. The latter was distributed between 500 ml CH 2 Cl 2 and 500 ml B 2 O, and the following sequence was repeated 3 times: The pH was adjusted with stirring to a constant pH of

2.5 with dilute HCl; The mixture was stirred vigorously for 25 minutes; and the H 2 O layer was separated, combined with 500 ml fresh CH 2 Cl 2, adjusted to a pH of

9.5 with dilute NaOH and the OHgOlg layer was separated.

The pH 9.5 CH2 Cl2 layer was combined with 500 ml fresh H2 O for sequence repetition. At a third stage, the pH 9.5 OHgOlg layer was dried (Na2 SO4) and processed to afford the crude title product as a foam, 34 g, which was crystallized from 150 ml of hot, cooled and diluted with 300 ml of pentane, yielding the purified title product,

25 »θ Si white crystals; dcd Rf 0.5 (CHCl 3: 9: 1: diethylamine); Rf 0.1 (CH2 Cl2 CH3 OH1 NH4 OH conc. 90: 10: 1), mp 170-18 ° C.⁰; 1 Hrmn (CDCl3) delta 2.26 / 6H, s, (CH2 N- ?, 3.29 (3H, s, cladinose CH2 O-); ppm 179.44 (lactone 0 = 0), 103.57 and 96.70 (0-3.0-5), 41.50 / (ch₃)₂-n-7

method Β

The title product of the previous non-chromatographed Example (6.2 g) was dissolved in 200 ml of ethanol and hydrogenated over 12.5 g of Raney Ni at 50 pounds per square inch for 18 hours. The reaction mixture was filtered, charged with 20 g of fresh Raney Ni and hydrogenation continued for 4 hours. Filtration and fresh catalyst refilling were repeated, and hydrogenation continued for a further 16 hours. Filtration and processing of the filtrate gave the crude title product as a white foam. The latter was partitioned between CH2 Cl2 and saturated NaHCO3, and the organic layer was separated, dried (1 SO4) and purified to give the title product as a second white foam, 5.6 g, crystallized as above. to provide the purified title product 955 mg, having identical physical properties to the product prepared by Method A.

EXAMPLE 8

4-Epi-9-deoxo-9a-hydroxy-9a-aza-9a-homo-erythromycin A 5 'N-oxide (X)

Shaking under N₂The title product of the previous Example (5.0 g) was dissolved in 15 ml of 1: 1: THE: CH3 OH. Thirty percent HgOg (5 ml) was added. After 0.5 hour, 50% H was added₂0₂ (5 ml). After a further 0.5 hour, the reaction mixture was cautiously soaked in 1: 1 CH 2 Cl 2 HgO containing excess Na 2 SO 4 (exothermic). The pH was 9. The aqueous layer was washed with CH₂C1₂ fresh and then with ethyl acetate. The organic layers were combined, dried (Na₂S0 ^) and process

<img file="PT77645B_D0026.tif" />

bags to provide the title product, 2.7 g ccf. Rf 0.15 (CHgClgzCH OHH OHHÍH ^OH conc. 60: 10: 1); ^ rmn (CDCl1)₅) delta 5.21 / δ6 s, s, (CH3 N7, 5.38 (3H, s, cladinosis CH2 O-); MS: higher peaks am / e 576 (ion of desosamine fragmentation at 0-5 ), 418 N-hydroxyglycone ion minus both sugars). Both peaks make a diagnosis for half -Ν-0Ξ with aglycone.

910 9

Soxo-9α-methyl-9α-aza-9α-homoerythromycin A 4-Epi-9-α^t-NH-0xIC (XI) title product from the previous Example (2.6 g 0.0054 mol) was dissolved in 100 ml CH₂C1₂ and strong stirring, K was added₂^Ç3 (37.5 g, 0.271 mol) and then CH3 (19.3 g 8.5 ml 0.156 mol) and the mixture was stirred for 20 hours. Filtration and work up gave the title product as a foam, 2.9 g tlc Ef 0.5 (CH₂C1₂: CH3 OH: NH4 OH conc. 60: 10: 1), Eph 0.15 (CH₂C1₂:: CH3 OH: NH3 OH conc. 90: 10: 1).

The title product prepared in this manner (2.8 g) was then purified by chromatography on 85 g of silica gel using CHgCEL ·,: CH ^OH: NH ^OH conc. 90: 10: 1. NH 4 OH as eluent; thereby removing minor impurities and polar impurities. Recovery: 0.87 gmHrmn (CDCl3) delta 2.52 (5 ', s, CH3 -N- aglycone), 3.20 /' 6 ', s, (CH3)₂ N,&gt; 07, 3.37 (3H, s, cladinosis OH 2 O-).

<img file="PT77645B_D0027.tif" />

ΕΧΕΜΡ10 1Q

4-Epi-9-deoxo-9a-methyl-9a-aza-9ahemoerythromycin A (IV)

Method A

Title product of Example (δ, 7θδ g, 0.96 mmol) was dissolved in 20 mL of CHCl3. Formaldehyde (57%, 0.078 mL) and then formic acid (0.03 mL) were added and the mixture was stirred for 4 hours, then refluxed for 7 hours. The reaction mixture. The reaction mixture was cooled, added to 30 ml of H₂0 and adjusted to pH 9 with 6N NaOH. The organic layer was separated, dried (NagSO4) and processed to afford the title product as a white foam, 0.7 g, crystallized from hot ethanol / H2 O, 302 mg, mp 153 °, recrystallized. from hot ethanol / H2 O, 246 mg; mp 155 °? Rf 0.55, (CH2 Cl2 CH3 OH: NE4 OH conc. 60: 10: 1) Rf 0.6 (CHCl3: diethylamine 9: 1); 1 Hrmn (CDCl 3) delta 2.29 ~ 9H, amplified, N-CH aglycone, and deosamine (CH 3)₅N, 7.31 (3Ξ, s, cladinosis CH2 O-); _{C; ra] n} (Qpcp, standard internal ObOlj) ppm 178.89 (lactone 0 = 0), 102.6% and 95.15 (C -3.0-5), 40.38 / - (CH2 N3; MS: peaks higher m / e 590 (N-methyl aglycone-ion desosamine via Cl cladinose cleavage), 416 /^_N-methyl ion aglycone (minus both sugars via Cl, C-5 cleavage) 7,158 (base peak, desosaminase-derived fragment).

Method E Unchromatographed title product from the title of the previous Example (0.242 g) and 10% Pd / C (0.4 g) were combined in 15 ml of 95% ethanol and the mixture was hydrogenated at 50 pounds per square inch. for 1 hour. 0 The catalyst was recovered by filtration and the filtrate was evaporated to afford the title product as a white foam, 160 mg, crystallized from ether / pentane, 124 mg recrystallized from ethanol / H2 O, 95. mg having the same physical properties as the title product by Method A.

<img file="PT77645B_D0028.tif" />

Method The title product of the previous Example chromatographically purified (319 mg) and Raney nickel (1.5 g wet-5% water) were combined in 20 ml ethanol and hydrogenated at 5θ pounds per square inch çtoantevl, 5 hours . The catalyst was removed by filtration and the mother liquor was evaporated to dryness to afford 205 mg of the title product, identical in physical properties to the title product by Method A.

EXAMPLE 11

2'-0-Acetyl-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin The title product of Preparation 5 (2.5 g 3) 34 mmol) was stirred with acetic anhydride (0.339 mL, 3). θθ mmoles) in 3 ° ml CH₂C1₂ for 4 hours.

The reaction mixture was purified and the residue was dissolved in 50 mL of ethyl acetate, combined with 50 mL of H2 O and the pH adjusted to 9) with 1 N NaOH. The aqueous layer was separated and washed with 20 ml of fresh ethyl acetate. The organic layers were combined, dried (NaSO4), processed, dissolved in 30 mL of CHCl3 and further purified to give the title product as a dry solid, 2.82 g, 2 Hrmn / CDCl3 including delta. 3.31 (C4-0CH₅), 2.28 (N-CH₅), 2,25 ₂J and 2.0 (2 * -0C0CH₅).

<img file="PT77645B_D0029.tif" />

EXAMPLE 12

2'-O-Acetyl-4-soxo-4-oxo-9-soxo-9a-methyl-9a-aza-9a-homoerythromycin A (VHIa)

The title product of the previous Example (2.5 g 3.2 mmol) and DMSO (0.38 mL 5.23 mmol) were dissolved in 9 mL of CH₂C1₂ and cooled to -70 ° C. Keeping the temperature below -50 ° C, trifluoroacetic anhydride (0.72 ml 4.95 mmol) was added by syringe and the mixture was stirred for 50 minutes at -60 ° C. Triethylamine (1.54 ml 11 mmol) was added by syringe, keeping the temperature below -50 ° C during the addition. The mixture was then warmed to 0 ° diluted with H2 O and the pH adjusted to 9.5 with diluted NaOH. The organic layer was separated, dried (NaSO 4) to afford the title product as a 2.5 g foam. The foam was flash chromatographed on silica gel with 10: 1 CHCl 3: CH 2 OH as eluent, monitored by tlc and collected 3 fractions. The purest fraction 1 of the product 1.7 g was dissolved in CHCl 3, diluted with B 2 O, adjusted to pH 4 with diluted HCl, and the aqueous layer was separated, diluted with fresh CHCl 3, adjusting. The pH was adjusted to 8 with dilute NaOH and the organic layer was separated.

The last aqueous layer was extracted with three portions of fresh CHCl3. The last four organic layers were combined, washed with H2 O, dried (N2 SO4) and processed to afford the purified title product.

0.98 g; Rf 0.7 (5: 1: 0.1 CHCl 3: CH 2 OH 3 OH 4 OH);¹Hrmn (CDCl3) includes delta (ppm), 2.05 (s, 3H, COCH3), 2.26 / s, 6H, N (OH₃)₂J, 2.33 (d, 3H, NOH) and 3.33 (d, 3H, OCH_?).

EXAMPLE 13

4-Lesoxy-4-oxo-9-deoxo-9a-methyl-9a-aaa-9a-homoerythromycin A (Vila) title product from previous Example (0.93 S s) dissolved in methanol. After 20 minutes the mixture was purified to afford the product of this title, 0.74 g; Nos. 746.4, 588.4, 575.4, 413.3, 158.1, 125.1; '-Ηπιιη (0D01₅) includes delta (ppm); 5.5 (t, 1H, 01-H), 4.6 (q, 1H, C5'-H), 3.35 (_s, 3H, OCH3), 2.38 (s, 3Ξ, NCH₅), 2.30 / s, 6H, N (CH₅)₂_7.

EXAMPLE 14

4-Epi-9-deoxo-9a-methyl-9a-aza-9a-homoerythromycin A (IV) title product of the previous Example (0.25 g) and Raney nickel 250 mg were combined in 20 ml ethanol and hydrogenated at 5θ pounds per square inch for 4 hours. The catalyst was removed by filtration and the filtrate was processed to an oil which crystallized on standing. 0 The title product was recovered by trituration with isopropyl ether and 0.13 g filtration, with identical properties to that of the Example product.

<img file="PT77645B_D0030.tif" />

PREPARATION 1

4-Epi erythromycin A

A suspension of 100 g Raney nickel paste in 1 liter of absolute ethanol containing 100 g of 4-deoxy-4-oxoerythromycin A (US 4,510,220) was stirred in a hydrogen atmosphere overnight at room temperature at 50 pounds. per square inch. The spent catalyst was filtered through the diatomaceous earth and the filtrate was concentrated in vacuo to 500 ml. Water (700 ml) was added to the concentrated filtrate and the resulting milky solution was heated on a steam bath. One piece added. a small amount of steam to prevent the transformation of the pro. gum duct upon precipitating from the solution. After stirring for 2 hours at room temperature the product was filtered and dried, 57.6 g and the filtrate was concentrated in vacuo to the point of nebrin. The mixture was allowed to stir for one hour and was filtered and dried, 21.4 g.

The resulting crops combined, 141-144 ° C. 1 H nmr (CDCl 3) showed absorption at

5.5 (5Ξ, s), 2.5 (6H, s) and 1.4 (5H, s) ppm.

PREPARATION 2

Erythromycin A Hydrochloride Oxime

Under, erythromycin A (500 g,

0.681 mole) was dissolved in pyridine (2.777 kg, 2.8501, 55.29 mole). Hydroxylamine hydrochloride (1.185 kg, 17.02 mol) was added and the mixture was stirred for 22 hours, then processed to a thick paste.

<img file="PT77645B_D0031.tif" />

and filtered with isopropanol as a wash. The combined filter with water was reprocessed to a thick, wax-like mass which crystallized by trituration with water, 615 g (slightly moistened with water, used in the next step without careful drying). ); ccd Rf 0.45 (CH₂C1₂: CH3 OH: NB3 OH conc. 60: 10: 1).

By the same procedure, 5 S of erythromycin A was converted to the dry title product, 4.5 g of at least 95% purity by NaCl. Recrystallization of 1 g from 10 ml of methanol and 10 ml of isopropyl ether gave 725 mg; mp 187 ° (dec) 157-160, 1970 bibliography o 15

188-191, Massey et al., Tetrahedron Letters, p. Crmnj / ~ DMSO- dg, ( CS4 )₄ internal standard Si7ppm 174.35 (lactone 0 = 0), 168.78 (G = N), 101.0 and 95.46 (C-3, C-5).

PREPARATION 3

9a-Aza-9a-homoerythromycin A

By the process of Example 2, with gas evolution verified when bicarbonate was added, with slight water humidification, the title product of the previous Preparation (615 S, evaluated to be 506 g, 0.613 mole on a dry basis was converted to 1 Crmn / CDCl3, internal standard GDCl7.7ppni 177.54 (lactone C = 0), 163.76 (amide 0 = 0 102.28 and 94.20 (C-3, C-5), 40.13 / (CH ^) ^ /

<img file="PT77645B_D0032.tif" />

PREPARATION 4

9-Deoxo-9a-aza-9a-homoerythromycin A

By reduction with NaBH 3 according to the method of Kobrehel et al. (supra) the title product of the previous preparation was converted to the title product.

PREPARATION 5

9-Deoxo-9a-methyl-9a-aza-9a-homoerythromycin A

By the procedure of Example 10 above, the title product of the above Preparation (21.1 g 0.028 µmoles) was converted to the product of this title, initially isolated as a white foam, crystallizing from hot ethanol / E2. 0.18 g, mp 136 ° C.