DK143032B - 7-POSITION PROTECTED 3-BROMETHYL-DELTA2-CEPHALOSPORANIC ACID ESTERS USED AS INTERMEDIATES IN THE PREPARATION OF DELTA3-CEPHALOSPORANIC ACID DERIVATIVES AND PROCEDURES FOR PREPARATION - Google Patents

Description

(\ fca> di) PUBLICATION NOTICE 1 ^ 3032 DENMARK (51), nt · C | · 3 c 07 D 501/60 uMIn IVInn IV C 07 D 601/04 § (21) Application No. 52 ^ 9/71 (22) ) Filed on July 1st. 1971 (24) Race day 6 Feb 19 (44) The application presented and the petition published on 16. 1 9 & 1

DIRECTORATE OF

PATENT AND TRADEMARKET SYSTEM (30) Priority requested from it

Feb 7 1968, 705525, US Jan 15 1969, 790842, US Jan. 15 1969, 790886, US

(71) ELI LILLY AND COMPANY, 507 East McCarty Street, Indianapolis, In = diana, US.

(72) Inventor: John Alan Webber, 642 Turtle Creek North Drive, Indiana Police, Indiana, US: Harle Marvin van Heyningen, 6020 East Raymond Street, Indianapolis, Indiana, US.

(74) Plenipotentiary in the proceedings:

Hofman-Bang & Boutard Engineering Company.

(54) at the 7-position protected 3-bromomethyl-delta2-cephalosporanoic acid esters for use as intermediates in the preparation of delta5-eephalosporanoic acid derivatives and process for their preparation.

The invention relates to novel 3-position protected 3-bromomethyl-Δ? -Cephalosporanoic acid esters for use as intermediates in the preparation of β-cephalosporanoic acid derivatives having antibiotic activity, which compounds are characterized by the formula of claim 1, and a process for the preparation of these compounds, which is characterized by the characterizing part of claim 2.

The semi-synthetic production of 7-acylamidodesacetoxycephalosporin antibiotics from penicillin starting materials has become important recently due to the method invented by Morin and Jackson (U.S. Patent No. 3,275,636) to convert penicillin sulfoxide esters for desacetoxycephalosporanoic acid esters. Cephalosporin compounds derived from penicillins in this process have the general formula 0 R-C-HN-CH-CH 2 1 2 CHO t t o j 2 CO-N 2 Jc-ch, 8 \ 3

T

COOR1 wherein R is the remainder of the acylamido group at the 7-position and R · 1- is hydrogen, a salt-forming cation, an ester group or anionic charge when C0000 forms a salt with a cation either inside or outside the molecule.

In the efforts to improve and extend the properties and utility of these penicillin-derived semi-synthetic cephalosporins, great efforts have been made to change the 3-methyl group of the above Δ ^ -desacetoxycephalosporins to a group which gives the resulting cephalosporin compound increased antibiotic activity. Gram-positive or Gram-negative microorganisms. However, to date, it has not been possible to directly convert an A 2 -desacetoxycephalosporin to a 3-methyl-derived A 3 ~ cephalosporin in any significant yield. Therefore, in the field of antibiotics, another method or pathway is needed to prepare the more effective Aβ-3-methyl-derived cephalosporin antibiotics obtained so far only from cephalosporin C produced by fermentation and the one obtained therefrom. obtained 7-aminocephalosporanoic acid (7-ACA).

The novel compounds of the invention are useful as intermediates in a chemical process, thereby circumventing the difficulty of converting -desacetoxycephalosporins into 3-methyl-functionalized β-cephalosporanoic acid antibiotics with a variety of substituents at the 3-position. It is surprising in itself that, after displacement of the double bond in Δ -desacetoxycephalosporins, these novel 3-bromomethyl-2, 143032 3 Δ-cephalosporanoic acid esters can be obtained, the bromine is replaced with a functional group and then the double bond is moved back to A position. , -z.

since direct bromination of the 3-methyl group in Δ -desacetoxy-cephaloranoic acid esters is not possible.

The 2-bromomethyl-β-cephalosporanoic acid ester intermediate of the invention can be reacted with a nucleophilic substance to replace the bromine from the 3-bromomethyl intermediate with the nucleophilic group 2 to form a 3- "functionalized-methyl" - / - cephalosporin ester product. The term "nucleophilic substance" in this specification is intended to mean a substance which gives a negatively charged group or neutral molecule bearing an undivided electron pair and which is part of a 2 nucleophilic substitution reaction with the 3-bromomethyl-A-cephalosporinester intermediate to form of a 3- (nucleophil methyl) -? A cephalosporine ester product. Numerous examples of nucleophilic substances that can be used are already known in the cephalosporin antibiotic patent literature for forming cephalosporin compounds with the nucleophilic group on the methyl group at the 3-position.

The 2β-7-acylamido-3- (nucleophil-methyl) -cephalosporine ester products can be partially converted to the corresponding Aβ-7-acylamido-3- (nucleophil-methyl) sulfide esters by heating in a weak basic medium to give the double bond to be moved to A ^ position to give an equilibrium mixture of the A ^ product and the Aβ-7-acylamido-3- (nucleophilic, methyl) cephalosporin ester, which is readily converted into an active antibiotic by removal of the ester group according to known methods, which are illustrated below. However, the 7-acylamido-3- (nucleophil methyl) -3-cephem-4-carboxylate ester product can also be chemically converted to the corresponding Δ -sulfide ester product of the corresponding Δ-sulfoxide ester with a peracid, (2) reduction of the 43-sulfoxide ester with a reducing agent such as sodium hydrogen sulfite or sodium dithionite in the presence of an activator such as acetyl chloride, in an organic solvent such as acetic acid or dimethylformamide the β-sulfide ester and (3), if desired, deesterification of the A ^ sulfide ester to the antibiotically active Δ · sulfide acid. If desired, mixtures of the active A-cephalosporanoic acid antibiotics and inactive Λ-cephalosporic acid and pharmaceutically acceptable salts thereof may be used in the mixed form for some antibiotic purposes, e.g. as topical antibiotic for open, wounds in veterinary medicine, in which case the mixture can be powdered on the wound or compounded in an ointment and applied in this medium to inhibit the growth of various Gram-positive or Gram-negative microorganisms.

The 2-desacetoxycephalosporin ester starting materials for the process of the invention can be obtained from a variety of penicillin or cephalo-sporin sources by known methods. They are obtained by treating the corresponding 3-methyl-A-cephem-4-carboxylic acid ester with base as described e.g. in Example 4 of U.S. Patent No. 3,275,626. They may also be obtained by hydrogenation of a cephalosporin C derived cephalosporin ester to form the corresponding Δ 2 -desacetoxycephalosporin ester and subsequent treatment thereof with a base such as pyridine in the cold (0 - 10 ° C) to isomerize the Δ ^ double bond to Δ2 position. Attempts to brominate the allylic 3-methyl group in the H 2 -desacetoxy-cephalosporanoic acid esters were unsuccessful. The successful bromination of the Δ -desactoxycephalosporanoic acid esters by the process of the invention was surprising and could not be predicted.

In the process of the invention, the esterified Δ 2-desacetoxycephalosporin starting material is mixed with N-bromosuccinimide in an organic liquid medium and stirred until the 3-bromomethyl product is formed. The reaction is preferably carried out in the presence of azobisisobutyronitrile as a catalyst or initiator to improve the yield, but the process can be carried out without the catalyst. The reaction takes place faster when the mixture is warmed slightly and for this reason the temperature range 40 - 100 ° C is indicated. The temperature must be kept below the decomposition point of the starting material and product. Also, too low temperatures cause the reaction to proceed too slowly for practical execution. The N-bromosuccinimide is usually present in at least stoichiometric amounts relative to the Δ -desacetoxycephalosporanoic acid ester, since the latter is the most expensive. The amount of azobisisobutyronitrile or other initiator used is not critical. As organic liquid diluents for use in the reaction mixture may be mentioned carbon tetrachloride or mixtures thereof with chloroform, tetrachloroethane, methylene chloride, benzene, toluene, xylene and heptane.

5 143032 2

Examples of the novel β-3-bromomethyl intermediates prepared by the process of the invention include: p-methoxybenzyl-7-phenoxyacetamido-3-bromomethyl-3-cephem-4-carboxylate o and tert, -butyl-7 phenoxyacetamido-3-bromomethyl-4-cephem-4-carbo-. xylate.

The invention is further illustrated by the following detailed examples, wherein Examples 1-5 illustrate the preparation of the intermediates of the invention and Examples 6-10 illustrate the use of these 2 for the preparation of 3-functionalized-methyl-4-cephalosporine ester products and their conversion to the corresponding to A 2 -cephalosporanic acids, which are active antibiotics.

EXAMPLE 1 2

Preparation of α-acid.

A portion of 3.63 g (0.01 mole) of methyl 7- (phenoxyacetamido) -3-methyl-β-cephem-4-carboxylate (prepared from desacetoxyceph V and diazo-methane) in 100 ml of pyridine / water (1: 1) was cooled in an ice-water bath.

1 equivalent of 1N NaOH solution was added and the mixture was stirred in the cold for 5 hours. After dilution with 100 ml of water and 100 ml of ethyl acetate, the mixture was cooled and acidified to pH 2.5 with 20% HCl. The ethyl acetate was removed and the aqueous layer was extracted once with ethyl acetate. The columnar organic layers were cooled, water was added and the pH was adjusted to 8.2 with solid NaHCO 3. The aqueous layer was separated, washed once with ethyl acetate, then cooled, placed under ethyl acetate and acidified to pH

The 2,5-ethyl acetate was removed, the aqueous layer washed with the ethyl acetate, and the combined organic layers washed twice with sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to a foam.

The foam product was dissolved in and crystallized from ethyl acetate to give 1.6 g (45 90 7- (phenoxyacetamido) -3-methyl-&-cephem-4-carboxylic acid, mp 180 - 183 ° C (d).

6 1A3032 o

Esterification of Λ -desacetoxycephalosnoranoic acid To a stirred suspension of 1.75 g (0.005 mol) of Δdesacetoxyceph V

p-7-phenoxyacetamido-3-methyl-ΰΓ-cephem-4-carboxylic acid and 700 mg (0.005 mole) of p-methoxybenzyl alcohol in 20 ml of methylene chloride were added a condensation solution consisting of 1.23 g (5% excess) of DMF dinopentyl acetate. methylene chloride. The solution was complete in minutes. The reaction mixture was stirred overnight at room temperature. Then, the solvent was removed and benzene was added. After heating to induce solution, the mixture was allowed to stand at room temperature.

After removal of a crystalline by-product, the benzene mother liquor was diluted, washed 3 times with hydrogen carbonate solution, 2 times with sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The residue was recrystallized from CCl ^ to give p-methoxybenzyl-7-phenoxyacetamido-3-methyl-Δ2-cephem-4-carboxylate. First crystallization gave 1.15 g, m.p. 108-112 ° C; second crystallization 0.16 g, m.p. 107 - 111 ° C; 55% yield.

Preparation of p-methoxybenzyl-3-bromomethyl-7-phenoxy-acetamido-β-cenhem-4-carboxylate

A mixture of 235 mg of p-methoxybenzyl-3-methyl-7-phenoxyacetamido-Λ and A. cephem-4-carboxylate, 90 mg of N-bromosuccinimide, 14 mg of azobis-isobutyronitrile, and 30 ml of CCll was heated at reflux under reflux. nitrogen and protected from light for 14 hours. The reaction mixture was cooled, filtered and the golden yellow filtrate evaporated to dryness. The structure of the crude p-methoxybenzyl-7-phenoxyacetamido-3-bromomethyl-2-cephem-4-carboxylate obtained was confirmed by NMR spectroscopy. The presence of the allylic bromide could be confirmed by maxima at cf 6.48 (roughly doubled) due to the vinyl hydrogen at the 2-carbon atom in the allyl bromide and at S 4.14 (quartet, J = 8 Hz) due to the methylene-bearing bromine atom. These absorptions infallibly show the presence of the allylic bromide in the crude reaction mixture.

EXAMPLE 2

Esterification.

2

To a stirred suspension of 6.98 g (0.002 mol) of 4-desacetoxy-ceph V-acid [3-methyl-7-phenoxyacetamido-β-cephem-4-carboxylic acid] and 2.8 g (0.002 mol) p-Methoxybenzyl alcohol in 100 ml of methylene chloride was added in solution of 4.62 g of dimethylformamide dinopentyl acetal in 25 ml of methylene chloride. The solution was complete in minutes. The reaction mixture was stirred for 48 hours at room temperature to ensure complete reaction. The solvent was removed and benzene was added. The resulting solution was washed three times with aqueous sodium bicarbonate solution, twice with aqueous sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The residue was recrystallized from carbon tetrachloride to give 6.74 g (92% yield of recovered β-acid) p-methoxy-benzyl-3-methyl-7-phenoxyacetamido-2-cephem-4-carboxylate, m.p. . 108-112 ° C.

The aqueous sodium bicarbonate mother liquors were shaken with ethyl acetate, cooled and adjusted to pH 2.5. The organic layer gave 1.56 g of colorless crystalline 3-methyl-7-phenoxyacetamido-A2-cephem-4-carboxylic acid which could be reused in the esterification process.

Functionalization of the 3-methyl group.

2

A portion of 1.17 g of p-methoxybenzyl-3-methyl-7-phenoxyacetamido-4, cephem-4-carboxylate was treated with N-bromosuccinimide in carbon tetrachloride in the presence of azobisisobutyronitrile to form p-methoxybenzyl.

ISLAND

benzyl 3-bromomethyl-7-phenoxyacetamido [beta] -cephem-4-carboxylate. The structure was confirmed by NMR.

EXAMPLE 3

Preparation of t-butyl 7-phenoyyacetamido-5-bromomethyl & 2-cephem-4-carboxylate.

A solution of 8.08 g (20 mmol) of t-butyl 7- (phenoxyacetamido) -3-methyl-A2-cephem-4-carboxylate, 5> 0 g of N-bromosuccinimide and 50 mg of azobisisobutyronitrile in 800 ml of carbon tetrachloride was heated. under reflux under nitrogen until a negative starch-iodine test (5 hours) was obtained. The reaction mixture was cooled, the succinimide removed by filtration, and the solvent removed under reduced pressure to give 11.8 g of crude product. An NMR spectrum showed only tert.-butyl-7-phenoxyacetamido-3-bromomethyl / JP-cephem-4-carboxylate contaminated with traces of succinimide (yield over 90%).

The NMR spectrum showed the following maxima: 1.56 (s) t-butyl 4.23 δ (q) -CH 2 Br

4,576 (s) PhOCH2CO

5.20 <5 '(s) C-4 5.33 6 (d) C-6 5.73 & (q) C-7 6.52 6 (s) C-2 vinyl 6.8-7.9 S (multiplied) aromatic and NH; at 1.9 S, there was only a slight maximum, barely above the noise level, which indicated a negligible amount of starting material.

EXAMPLE 4 p

Preparation of nitrobenzyl 7-phenoxyacetamido-3-bromomethyl-4-cephem-4-carboxylate 2

A mixture of 4.835 g of p-nitrobenzyl-7-phenoxyacetamido-3-methyl-AT-cephem-4-carboxylate, 80 ml of chloroform and 320 ml of carbon tetrachloride was heated to dissolve the whole. To this solution was added 2.67 g of powdered N-bromosuccinimide and 82 mg of azobisisobutyronitrile and the mixture was kept in an oil bath at 85 ° C for 2 hours and 15 minutes. The reaction mixture was cooled to room temperature and the solvents evaporated. An aliquot was examined by NMR which confirmed the structure as the 3-bromomethyl compound.

The NMR spectrum showed the following maxima: 4.16 <P (q) -CH2Br 4.55 tP (s) -CH2- in 7-side chain 5.30 cT (s) -CH2- in ester 5.75 S (<l ) H 1 position 6.58 ^ (s) Hi 2 position 7.14 tT (multiplet) H on phenyl ring 7.58 and 8.15 oΓ (doublets) H on benzyl ester.

The following examples illustrate the use of the intermediates of the invention.

EXAMPLE 5

Functionalization of p-methoxybenzyl-3-bromomethyl-7-phenoxyacetamido-A-cephem-4-carboxylate

To the crude product of Example 1 was added 25 mg of potassium acetate and 15 ml of acetone, and this mixture was heated at reflux for 6 hours under nitrogen protected from light and then stirred at room temperature overnight. The dark reddish brown solution was evaporated to dryness, the residue taken up in chloroform, filtered and evaporated to give 250 mg of a reddish brown oil.

This oil was purified twice by pisseparative thin layer chromatography, 2 hours to obtain 103 mg of a mixture of the Δ - and Δ-β-methoxy benzyl ester of ceph V.

2

Cleavage of the mixed Δ -Δ ester product with trifluoroacetic acid in benzene gave a material containing ceph V acid, a known anti-biotic, as shown by thin layer chromatography and a bioautogram of a paper chromatogram.

2 3

Oxidation of the ester mixture with m-chloroperbenzoic acid in chloroform yielded a high yield of the corresponding p-methoxybenzyl-3-acetoxy-methyl-7- (2'-phenoxyacetamido) -A 2 -cephem-4-carboxylate -1 oxide, mp.

161 - 163 ° C (from methanol).

Reduction of p-methoxybenzyl-3-acetoxymethyl-7-phenoxyacetamldo-Δ-cephem-4-carboxylate-1-oxide

To a solution of 500 mg of the sulfoxide, p-methoxybenzyl-3-acetoxy-methyl-7- (2'-phenoxyacetamido) -A 2 -cephem-4-carboxylate-1-oxide, in 40 ml of dimethylformamide was added 10 ml of acetyl chloride and then 3 g of sodium diethionite.

There was an exothermic reaction. After stirring for 4 hours at room temperature to ensure complete reaction, the mixture was cooled, diluted with benzene and neutralized with aqueous sodium hydrogen carbonate. After the vigorous gas evolution died out, more benzene was added and the organic layer was separated and washed thoroughly with aqueous sodium hydrogen carbonate solution and then with sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 635 mg of brown oil.

This product was purified on a column of silica gel with 15% H2 O. 253 mg of an oil were obtained whose NMR, IR and UV spectra were the same as those of p-methoxybenzyl 7- (2 H -phenoxyacetamide) -cephem-4-carboxylate prepared from 7-amino This oily ester product could be crystallized from ether to give 195 mg of material, mp 118 - 119 ° C, the melting point of which was not lowered when mixed with samples of the known compound.

Cleavage of p-methoxybenzyl ester to ceph V

To a solution of 105 mg of p-methoxybenzy 1-7-phenoxyacetamido-3-acetoxymethyl-β-cephem-4-carboxylate and 22 mg of anisole in 10 ml of dry benzene was added 0.5 ml of trifluoroacetic acid. The mixture was stirred for two hours at room temperature under nitrogen and then evaporated to dryness. The residue was taken up in ethyl acetate and extracted three times with aqueous sodium hydrogen arbonate solution.

The ethyl acetate solution was evaporated to give 55 mg of neutral product.

The hydrogen carbonate extracts were cooled, shaken with ethyl acetate and adjusted to pH 2.4 by 20 pet. HC1. The ethyl acetate layer was separated and the aqueous portion was extracted once with ethyl acetate. The combined organic layers were washed twice with sodium chloride solution, dried, filtered and evaporated to give 86 mg of a colorless oil having the same R value as 7-phenoxy-2 L acetamido-3 acetoxymethyl-A-cephem-4-carboxylic acid in a thin layer chromatogram. The chloroform solution of this oil precipitated a solid which was removed by filtration. The filtrate was evaporated to give 65 mg of product which was found to be 7-phenoxyacetamido-3- 2 acetoxymethyl-4-cephem-4-carboxylic acid, since its NMR spectrum was identical to that of a known sample of this compound, a known antibiotic, which until now could only be obtained from fermented 7-aminocephalosporanoic acid.

EXAMPLE 6 11 143032 2 3

Preparation of methyl 3-acetoxymethyl-7-phenoxyacetamido-Δ - and Δ-cephem-4-carboxylate mixture.

2

To 110 mg of methyl 3-bromomethyl-7-phenoxyacetamido-n-cephem-4-carboxylate were added 25 mg (0.25 mmol) of potassium acetate and 15 ml of acetone. The mixture was heated under reflux under nitrogen and protected from light for 6 hours and stirred at room temperature overnight. After the solution was evaporated to dryness, the residue was taken up in chloroform and filtered. Evaporation of the solvent gave 130 mg of a golden oil.

Separation of the oil into its components by preparative thin layer chromatography yielded 11 mg of recovered starting material and 70 mg (65 pct. Yield) of a mixture of methyl 3-acetoxymethyl-7-phenoxyacetamido-4β-cephem-4-carboxylate (the structure confirmed by NMR; ratio 3: 1).

EXAMPLE 7

Preparation of methyl 3-azidomethyl-7-phenoxyacetamidoceph-Δ2 and c ephem-4-carboxylate mixture o

To 110 mg of methyl 3-bromomethyl-7-phenoxyacetamido-4-cephem-4-carboxylate were added 16 mg of sodium azide (0.25 mmol) and 15 ml of acetone. The mixture was heated at reflux for 6 hours and then stirred overnight at room temperature. The acetone was evaporated and the residue taken up in chloroform and filtered. Removal of the solvent gave 123 mg of brown oil.

The brown oily crude product was purified by preparative thin layer chromatography. In this way, 79 mg of a material containing both starting material and the azido product was obtained (shown by I.R. spectrum). Examination by NMR and IR revealed that this material contained 16 per cent. starting material (1: 1, Δ-year) plus the desired mixture of methyl 3-azidomethyl-7-phenoxyacetamido-β-cephem-4-carboxylate and methyl 3-azidomethyl-7-phenoxyacetamido - β-cephem-4-carboxylate (3: 1 & 2-A ^) in 65% yield.

EXAMPLE 8 p-Methoxybenzyl-3-bromomethyl-7-phenoxyacetamido-A-cephem-4-carboxylate product of Example 2 (1.25 g) was dissolved in 100 ml of absolute methanol containing 2 molar equivalents Ν, Ν- diethylaniline to absorb the hydrogen bromide by-product. This mixture was stirred at room temperature for 24 hours to ensure complete reaction and then evaporated to dryness, taken up in benzene, extracted twice with cold 5% hydrochloric acid and with sodium hydrogencarbonate solution and sodium chloride solution, dried over magnesium sulfate, filtered and evaporated to give 970 mg of a brown oil. This brown oil was chromatographed over a column of silica gel containing 15% water, using a mixture of benzene and ethyl acetate as the eluent.

A benzene solution containing approx. 4% ethyl acetate eluted ca. 15% low reacted desacetoxy starting material. A benzene solution containing approx. 8% ethyl acetate eluted the p-methoxybenzyl-3-methoxymethyl-7- Λphenoxyacetamido-Δ-cephem-4-carboxylate ester, m.p. 116-118 ° C from methanol, in 40% yield. The structure was confirmed by NMR spectrum and elemental analysis.

sulfoxide

To a cooled solution of 215 mg of p-methoxybenzyl-3-methoxymethyl-7-phenoxyacetamido-A2-cephem-4-carboxylate in 5 ml of chloroform was added a solution of 85 mg of m-chloroperbenzoic acid (88% pure) in chloroform. The mixture was stirred and allowed to warm slowly over one hour. The solution was washed with sodium bicarbonate solution and sodium chloride solution, dried over magnesium sulfate, filtered and evaporated. The solid residue was recrystallized from benzene containing methylene chloride to give 82 mg of p-methoxybenzyl-3-methoxymethyl-7-phenoxyacetamido - ^ - cephem-4-carboxylate-1-oxide, m.p.

183 - 185 ° C which gave a satisfactory elemental analysis.

Further experiments have shown that hydroxyl solvents such as isopropanol or tert-butanol yield better yields than chloroform in this reaction.

Reduction of sulfoxide.

To a solution of 1.028 g (0.002 mol) of p-methoxybenzyl-3-methoxymethyl-1-7-phenoxyacetamido-α-cephem-4-carboxylate-1-oxide in 75 ml of dry 13 was added 15 ml of acetyl chloride and then 6 g of sodium diethionite (Na 2 S 2 O). After stirring at room temperature for 4 hours, the resulting mixture was cooled and benzene and aqueous sodium hydrogen carbonate solution were added. After the decomposition of the excess acetyl chloride was complete, water was added and the mixture was extracted twice with benzene. The combined benzene extracts were washed with aqueous sodium hydrogen carbonate solution and with sodium chloride solution, then dried over magnesium sulfate, filtered and evaporated to give 1.32 g of crude, dark brown, semi-solid p-methoxybenzyl-3-methoxymethyl-7-phenoxyacetamido ^ -cephem-4-carbo-xylate.

This raw Δ. Cephalosporin ester was purified by column chromatography and eluted from a column of silica gel containing 15% water using benzene with 5% ethyl acetate as eluant. 300 mg of p-methoxybenzyl-3-methoxymethyl-7-phenoxyacetamido-h-cephem-4-carboxylate were obtained, m.p. 116 - 117.5 ° C after crystallization from ethyl ether. The assumed structure was confirmed by NMR spectrum and elemental analysis.

Esterf.iemelse

To a solution of 174 mg of p-methoxybenzyl-3-methoxymethyl-7-phenoxy-acetamido-Δ-cephem-4-carboxylate and 119 mg of anisole in 25 ml of dry benzene is added 1.25 ml of trifluoroacetic acid. After stirring for 2 hours at room temperature, the reaction mixture was evaporated, the residue was taken up in ethyl acetate and the resulting ethyl acetate mixture was extracted three times with aqueous sodium hydrogen carbonate solution.

The aqueous hydrogen carbonate extracts were cooled, shaken with ethyl acetate and acidified to pH 2.8. Evaporation was obtained from the ethyl acetate layer by evaporation of 140 mg of foam giving 86 mg of 3-methoxymethyl-7-phenoxyacetamido-1β-cephem-4-carboxylic acid, m.p. 135 - 137 ° C, after recrystallization from ether. This acid gave an antibiotic active zone on a bioautogram of its paper chromatogram. The stain for the derivative moved slightly slower than the stain for desacetoxyceph V.

EXAMPLE 9

Tert.-Hutyl-7-phenoxyacetamido-3-isopropoxy methyl-2

A solution of 3> 51 g tart. -butyl 7-phenoxyacetamido-3-bromomethyl-2 2-cephem-4-carboxylate, prepared as described in Example 3, in 100 ml of isopropanol was heated on a steam bath to ca. 70 ° C for 20 minutes. The excess isopropanol was removed and the pressure decreased. The residue was dissolved in benzene, washed with aqueous 5 pct. sodium bicarbonate solution and with water, dried over sodium sulfate and evaporated to dryness to give 2.32 g of crude tert.-butyl-7-phenoxyacetamldo-3-isopropoxymethyl-α-cephem-4-carboxylate as product. Spectral analyzes confirmed the structure.

Oxidation o 2.32 g of crude tert.-butyl-7-phenoxyacetamido-3-isopropoxymethyl, 0-cephem-4-carboxylate was dissolved in a mixture of methylene chloride and 50 ml of isopropanol and stirred at ice temperature while a solution was added dropwise. of 1.00 g of m-chloroperbenzoic acid in 50 ml of methylene chloride. Upon completion of the addition, the solvent was removed under reduced pressure. The residue was dissolved in ethyl acetate, washed with aqueous 5% sodium hydrogen carbonate solution, dried over sodium sulfate and evaporated to dryness to give 2.16 g of the crude sulfoxide ester, tert-butyl-7-phenoxyacetamido-3-isopropoxy-methyl N, -cephem-4-carboxylate-l-oxide. The crude sulfoxide ester was chromatographed over 200 g of silica gel containing 15% water to give 1.10 g of the pure sulfoxide ester in a mixture of 40% ethyl acetate and 60% benzene. The structure of the pure sulfoxide ester was confirmed by spectral data.

Reduction-

A solution of 0.891 g of the sulfoxide ester, tert.-butyl-7-phenoxyacetamido-3-isopropoxymethyl - ^ - cephem-4-carboxylate-1-oxide, 0.71 g of stannous chloride, 0.5 ml of acetyl chloride, ml of acetonitrile and 5 ml of dimethylformamide were stirred at 25 ° C for 2 hours. The solvents were removed under reduced pressure and the residue was suspended in ethyl acetate. The ethyl acetate suspension was washed twice with aqueous 3% hydrochloric acid solution and with aqueous 5% sodium bicarbonate solution and then with water, dried over sodium sulfate and evaporated to dryness to give 0.781 g of the reduced ester (sulfide), tert. -butyl 7-phenoxyacetamido-3-isopropoxymethyl-A ^ -cephem-4-carboxylate. Spectral analyzes confirmed the structure.

5 143032

deesterification

A solution of 0.156 g of tert.-butyl-7-phenoxyacetamido-3-isopropoxy-methyl-α-cephem-4-carboxylate in 7 ml of 98 pct. formic acid and water were stirred at 25 ° C for 25 hours to ensure complete reaction. The reaction mixture was poured into water and extracted with ethyl acetate. The ethyl acetate solution was washed with water and then extracted with aqueous 5 pct. sodium. The resulting solution was acidified with aqueous 5 pcts. hydrochloric acid and extracted with ethyl acetate. The resulting ethyl acetate solution was dried over sodium sulfate and evaporated to dryness to give 0.088 g of the free acid, 7-phenoxyacetamido-3-isopropoxymethyl - staphylococci by in vitro experiments.