SE466203B - ESTRARS FOR PREPARING A BETA-LACTAMAS INHIBITOR - Google Patents

Description

466 203 2 The formation with the structure s a S a¿N .__ å__š ° H2 ° 1 (IH: // 'ä.

Q CCXDH is disclosed in U.S. Patents 3,954,732, 3,989,685, 3,993,646, 4,009,159 and 4,036,847.

U.S. Patent No. 4,155,912 discloses 2-penem-3-carboxylic acid compounds having the form Ffs / 'B 4-N CCKDH' and esters and salts thereof; see also Farmdoc 8209UA, T0336B and 44337BJ The compound (designated CP-45899) having the structure coon is an irreversible beta-lactamase inhibitor with excellent stability in solution. It has weak antibacterial activity and potentiates the in vitro and in vivo activity of ampicillin with respect to beta-1-lactamase-producing strains [Ã.R. English §§_al: Antimicrobiaï Agents and Che- motherapy_l fi (1978) '414-419; Aswapokee gt_al .: J. Antibiotics §l1l§l De. 1978) 1238-1244; Derwent's Farmdoc Abstracts 89627A and 7386657. 466 2Û3 By B. Ba1tzer et al. combining beta-lactam antibiotics with a beta-lactamase inhibitor in a single molecule, which serves as a prodrug for the two active components, can be impaired by the coupled esters 3 and 4 listed below, in which ampicillin and meci11inam, respectively, are combined with beta-1actamase inhibitor penici11ansyrasu1fon. The authors show that these esters in humans are extremely well absorbed from the gastrointestinal tract and, after absorption, are hydrolyzed while simultaneously releasing the active components. As a result, high blood and tissue levels of antibiotic and beta-1-lactamase inhibitor are achieved in a balanced ratio. The benefits of “mutua1 pro-drugs” compared to single combinations are discussed.

The above-mentioned esters 3 and 4 have the structural forms © - fïï, '“co cH-co-Nim o NH ä Ek, o N ._ 0, ° o o * f _ o Rs? q / l / í / ~ / f- '-co -o -n -] _ o GÅ co- In GB 2 044 255 published on October 15, 1980, the hitherto unknown compounds of the general formula 1 are rejected: H Rl- cH-co-Nak; .

RQ. wherein R 1 represents phenyl, 4-hydroxyphenyl, 1,4-cyclohexadienyl or 3-thienyl, R 2 represents a primary amino or a carboxy group, R 3 represents a hydrogen atom or lower alkyl, aryl or aralkyl and A represents a beta-lactamase inhibitor group containing a beta-lactam ring as well as a carboxy group, wherein A is attached to the rest of the molecule via the carboxy group.

The new compounds are useful in the treatment of bacterial infections and are particularly active against beta-lactamase-producing bacteria. See also Farmdoc 6D773C and 607766.

Accordingly, the present invention provides the acid of formula O. \\ 4 * 0 a; \\ s cnzcl s' 11 ,, I I CH3 öâF__N_ Éoon and pharmaceutically "acceptable salts" and "Densely hydrated" esters "of the said acid.

The pharmaceutically acceptable salts contemplated in the present context include non-toxic metal salts such as salts of sodium, potassium, calcium and magnesium, ammonium salts and substituted ammonium salts, for example salts of such non-toxic amines as trialkylamines (eg triethylamine), procaine, dibenzylamine, N-benzyl-beta-phenethylamine, 1-ephenamine, N, N'-dibenzyl-ethylenediamine, dehydroabietylamine, N, N'-bis (dehydroabiet-ethyl) ethylenediamine, N- (lower alkyl) piperidine (for example N -ethylpiperidine) and other amines, which have previously been used for the preparation of pharmaceutically acceptable salts of penicillins and cephalosporins. The most preferred salts are the alkali metal salts, i.e. the sodium and potassium salts, and the ammonium salt.

By "easily hydrolyzed esters" in the present context is meant those pharmaceutically acceptable esters known in the art which are hydrolyzed to the free acid form in vivo. Examples of suitable easily hydrolyzed esters include phenacyl, acetoxymethyl, pivaloyloxymethyl, alpha-acetoxyethyl, alpha-acetoxybenzyl, alpha-pivaloyloxyethyl, phthalidyl (3-phthalidyl), indanyl (5-indanyl), methoxymethyloxyethylbenzyl methyl, propionyloxymethyl, valeryloxymethyl, isobutyryloxymethyl, 6-LKR) -2-amino-2-phenylacetamide-3,3,3-dimethyl-7-oxo-4-thia-1-azabicyclo [3.3.2] heptane-2-carbonyloxymethyl and 6 -LKR) -2-amino-2-p-hydroxyphenylacetamide--3,3-dimethyl-7-oxo-4-thia-1-azabicyclo13.2.Q] heptane-2-carbonyloxymethyl.

The preferred esters are acetoxymethyl, pivaloyloxymethyl, methoxymethyl, phthalidyl, 5-indanyl, 6-LR) -2-amino-2-phenylacetamide (7,3,3-dimethyl-7-oxo-4-thia-1-azabicycloL3). 2.Qheptane-2-carbonyloxymethyl and 6- [1R) -2-amino--2-p-hydroxyphenylacetamideQ7-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo [3.2.Q7- heptane-2 -carbonyloxymethyl.

Furthermore, according to the present invention there is provided a process for the preparation of the acid of the formula ffgfëlfl or a pharmaceutically acceptable salt or a slightly hydrolysed ester thereof, the process being characterized by a) catalytically hydrogenating, for example with a noble metal catalyst such as palladium, an ester of the formula i cxazcl .BI-bh. ï fil; wherein R 1 is benzyl or substituted benzyl and then b) oxidizes the hydrogenated product to produce the desired acid or salt thereof and then, if desired, c) esterifies the acid or the salt thereof to produce a readily hydrolyzed ester of the acid.

The present invention further provides a process for the preparation of the acid of the formula o o \ s / cnzcl I 'C33' _14 'z? E 10 H or a pharmaceutically acceptable salt or a slightly hydrolysed ester thereof, the process being characterized in that a) oxidizing, for example with KMnO 4, H 2 O 2 or similar peroxide or peracid, an ester of the formula O.i II ", rN 0 ä-0CHéCCl 3 for the preparation of an ester sulphoxide of the formula 0 0 Good 7 466 zoa and then b) reacting the ester sulphoxide with a metal in acid, such as with zinc in glacial acetic acid, to prepare the desired acid or salt thereof and thereafter, if desired ic) esterifies the acid or salt thereof to produce a lightly hydrolyzed ester of the acid.

A variety of oxidants known in the art for oxidizing sulfides to sulfones can be used. Particularly suitable reagents, however, are alkali metal permanganates, for example potassium permanganate, and organic peracids, for example 3-chloroperbenzoic acid.

Particularly useful protecting groups for the substituent R 1 are the benzyl group and substituted benzyl groups, especially 4-nitrobenzyl. The benzyl group and substituted benzyl groups can be conveniently removed by catalytic hydrogenation. In this case, a solution in an inert solvent of the compound of formula A above is stirred or shaken, wherein R 1 is benzylphenylated, substituted benzyl, under an atmosphere of hydrogen or hydrogen mixed with "an" inert diluent such as nitrogen. or argon in the presence of a catalytic amount of a hydrogenation catalyst. Suitable solvents for this hydrogenation are lower alkanols such as methanol, ethers such as tetrahydrofuran and dioxane, low molecular weight esters such as ethyl acetate and butyl acetate, water and mixtures of these solvents. However, it is common to choose conditions under which the starting material is soluble. The hydrogenation is usually carried out at a temperature in the range 0-60 ° C and at a pressure of 1-10 kg / cm 2. The catalysts used in this hydrogenation reaction belong to the type of agents known in the art for this type of reaction and suitable examples are noble metals such as nickel, palladium, platinum and rhodium. The catalyst is usually present in an amount of 0.01 to 2.5% by weight and preferably 0.1 to 1.0% by weight, based on the compound of formula A. It is often convenient to suspend the catalyst on an inert support; a particularly suitable catalyst is palladium deposited on an inert support such as carbon. In addition, it is common to buffer the reaction mixture in order to operate at a pH in the range of 4-9 and preferably 6-8.

Borate and phosphate buffers are commonly used. The reaction usually takes approx. 456 203 l time.

According to the invention there are further provided esters of the formula c R-QoH-co ma ... S Fä N22 I "C153 f _ I .0 / l '" c = ooo I _ é _ Q s * cnzcl (I) I,' _ |. I fç eng 4 "" * - zíä I .o c ---- o II, o varxïR-ä-r 'O. <:', F H2 enar 31 where R] is hydrogen or hydroxy and Rz is hydrogen , hydroxy, methyl, methoxy or chlorine.

The invention preferably relates to the esters of the formula 9 466 203 m2 ¿, __ N ___ '. ,, 1 H 3 O "c = o I l' ff C CH2 ár-'èlf. 1 ,, fi; I o - f; 0 and of the formula and also a process for the preparation of such an ester, which process is characterized by treating with acid a solution of a compound of the formula 'lf 0 C 0 || Û RNRWÖQÉ. Q: G, ES¶ II or _ H, where R4 is hydrogen or hydroxy and R5 is hydrogen, R. hydroxy, methylg methoxyyl ie chloro and vari "wherein * REräF" äTKyT; aratkyl * eTler * aryl, R2 is hydrogen, alkyl, aralkyl or aryl and R3 is alkyl, aralkyl, aryl, alkoxy, aralkoxy, aryloxy or R 11 -Nv 2, R 5 wherein R 4 and R 5 are each hydrogen, alkyl, aralkyl or aryl or together with the nitrogen atom represent piperidino or morpholino, the treatment with the acid being preferably carried out in an organic solvent such as acetone or chloroform or in aqueous solution or partial aqueous solution and preferably between pH 1 and pH 5 at room temperature.

It is further preferred that in the amino protecting group R 1 is methyl, R 2 is hydrogen and R 3 is methoxy, ethoxy or methyl; this requires the use of methyl acetoacetate, ethyl acetoacetate or acetylacetone.

When removing the alpha-amino protecting group, it is preferred to use a strong mineral acid such as hydrochloric acid or to use formic acid.

The present invention also provides as novel intermediates esters of the formula S C82Cl ff, 3: f "01 äs. Wherein R is benzyl or substituted benzyl, preferably p-nitrobenzyl, and a process for the preparation thereof, which process is characterized by heating , preferably under reflux, a compound having the formula B - O Bmx, -E fNv / 'in ink 0 /. * g-g3 ,, _ »0 wherein R is benzyl or substituted benzyl and preferably p-nitrobenzyl, in an inert, anhydrous organic solvent, preferably dioxane, in the presence of large and preferably equimolar amounts of a weak tertiary amine, preferably quinoline, and an acid chloride, preferably benzoyl chloride, until the reaction is substantially complete.

Furthermore, according to the invention there are provided as new intermediates esters of the formula ~; wherein R is benzyl or substituted benzyl, preferably p-nitrobenzyl, and a process for the preparation thereof, which process is characterized by oxidizing a solution in an inert solvent, preferably methylene chloride, of a compound of the formula Br Wherein R is benzyl or substituted benzyl, preferably p-nitrobenzyl, at approximately room temperature using a peracid, preferably m-chloroperoxybenzoic acid.

The present invention also provides as a novel intermediate the ester of the formula and a process for its preparation, which process is characterized by oxidizing a solution in an inert solvent, preferably methylene chloride, of a compound of the formula Jr I 2 caecl 'fo,; S At about room temperature by using an oxidizing agent such as KMnO 4, H 2 O 2 or similar peroxide or preferably a peracid, especially m-chloroperoxybenzoic acid.

B 466 203 The invention is further illustrated by the following working examples, in which the temperatures refer to degrees Celsius. "Skelïysoïve B" is a petroleum meter fraction with a boiling point of 60-68 ° C, which consists mainly of n-hexane.

Example 1 Preparation of potassium-2beta-chloromethyl-2a1fa-methylpenam-3a1fa-carboxylate-sulfone §BL-P2013) | »-. | aa BI 'å' efnuïš Qåï i àíf as? få 'à [W o \ oi CHZCI mo \ s7 cazcl' ff 4 š r- “I * fen Ye 3 Ä I 3 N _ N 2025 0 // l / 'cozx (BL-PZOI3) 466 203 M 6alpha-bromopenicillanic acid -S-sulfoxide (11l, 37.5 mmol of 6alpha-bromopenicillanic acid N, N'-dibenzylethylenediamine salt LG. Cignarella et al .: J.0rg. Chem. Gl (1962) 2668 and E. Evrard: Nature _2_0_l_ (l964 in 330 ml of methylene chloride, the mixture was stirred and cooled to 0 DEG C. Then 13 ml (156 mmol) of concentrated hydrochloric acid were slowly added to the methylene chloride solution, and the dibenzylethylenediamine HCl salt (DBED; HCl) precipitated within one minute. stirred for 10 minutes at 0-5 ° C.

To remove the DBED-HCl precipitate, the mixture was filtered through a diatomaceous earth filter ("Dicalite") and the filter cake was washed with 150 ml of methylene chloride. (Filtration should be completed as soon as possible and the acidic methylene chloride solution should be avoided for a long time. Some filtration problems may arise due to the fineness of the precipitate, so the addition of filter aids to the slurry may be helpful).

It: combined.methylene clonide meditrate was washed out with ka m fi fkaält fl vat fi emwf * while stirring for 5 minutes; after which the water phase was discarded. The pH of the washing liquid was 2.0-2.3. The methylene chloride solution containing 6alpha-bromopenicillanic acid was concentrated under reduced pressure to a volume of 65-80 ml.

The solution was cooled with stirring to 5 ° C.

While stirring vigorously, 13 ml (86.9 mmol) of 40% peracetic acid was carefully added over 30 minutes. The reaction was exothermic and the temperature was maintained between 15 ° C and 18 ° C by ice bath cooling. The sulfoxide began to crystallize out after the addition of 10 ml of peracetic acid. The slurry was cooled with stirring to 0-5 ° for 2 hours.

The slurry was filtered and the snow-white filter cake was washed as follows: 10 ml of water with a temperature of 5 °, then 10 ml of methylene chloride with a temperature of 0-50 and finally 15 ml of heptane.

The filter cake was dried in an oven at the temperature of 45 ° to constant weight, which took 6-10 hours. (Too prolonged drying can produce traces, 5,466,203 of a light red color). The amount of product 1 was 16.26 g and the yield 73.241.

The reaction mixture and the final product could be monitored by thin layer chromatography using the solvent systems 15 parts toluene / 4 parts acetone / 1 part acetic acid or 8 parts acetone / 8 parts methanol / 3 parts toluene / 1 part acetic acid. The final product was analyzed by NHR and IR. nitrobenzyl 6-alpha-bromopenicillanate S-sulfoxide (2).

To a solution of 12 g (0.04 mol) of 6alpha-bromopenicillanic acid S-sulfoxide in 100 ml of acetone was added 7.5 g (0.04 mol) of potassium 2-ethylhexanoate. The salt was collected by filtration, washed with cold acetone and air dried to give a total of 10 g. The crystalline potassium salt was dissolved in 75 ml of dimethylacetamide and 7.8 g (0.04 mol) of p-nitrobenzyl bromide were added. The solution was stirred for 24 hours at 23 °. The mixture was diluted with 500 ml of water and extracted with ethyl acetate. The ethyl acetate layer was washed 4 times with water and dried over anhydrous magnesium sulfate. The solvent was evaporated at 35 ° (15 mm) to an oil which crystallized. The light brown crystals of product 2 were slurried by ether ether to filtrate, filtering in a yield of 9 g (70%) and with a melting point of 24 DEG-24 DEG-25 DEG C. (Sun A distribution).

Analysis Ber. for C 15 H 15 BrN 2 O 6 S: C 41.98 H 3.05 N 6.52 Found: C 42.00 H 3.48 N 6.98 IR (KBr): 1800 (s), 1740 (s), 1610 ( w), lS20 (s), l450 (m), l350 (s), l060 (m), 740 (m) cm_]. 1 H-NMR (60 mHz, DMSO): δ 1.22 (s, 3H), 1.6 (s, 3H), 4.67 (S, 1H), 5.2 (d, Jnfl-5 Hz, 1H), 5.45 (s, 2H), 5.68 (d, Jfvl-5 Hz, 1H), 7.5-8.5 (m, 4H). nitrobenzyl 2-beta-chloromethyl-Zalfa-methyl-6-bromopenam-3alpha-carboxylate (§) A solution of 5 g (0,0l2 mol) of p-nitrobenzyl-6alpha-bromopenicillanate S- -sulfoxide (2) in 120 ml of anhydrous dioxane was refluxed for 4 hours under nitrogen with 1.5 g (0.012 mol) of quinoline and 1.6 g (0.012 mol) of benzoyl chloride.

The solution was diluted with 600 ml of water and extracted with ethyl acetate. The ethyl acetate extract was washed with a 5% sodium bicarbonate solution, a 466,203% W phosphoric acid solution and finally with water. The organic layer was dried over anhydrous magnesium sulfate and evaporated to an oil at 35 ° (15 mn). The ethanes were crystallized and taken up, washed with ether and finally with calcium toluene to give 3.5 g of the product 3 (ssz), m.p.

Ana1Xs Ber. for C 15 H 15 ClBrN 2 O 5 S: C 40.06, H 3.14 N 6.23 Found: C 40.19 H 3.12 N 6.75 1R (kßr) = 1792 (s), 174o (s), 1e10 (w) , 1520 (s), 1ssa (s), 1zso (m), 1o2s (w), 99o (w), 7so (w) cm -1, NMR (so mHz, nMso) = di 1.45 (s, 3H ), 3.5-4.3 (m, 2H), s, os (s, 1H), 5.42 (s, 2H), s, s (d, a-1.5 Hz, 1H), , 62 (d, a «« i, s Hz, 1H), 7, ss, s (m, 4H). p-Nitrobenzyl-Zbeta-chloromethyl-2α-alpha-methylpenam-6a1-alpha-carboxylate sulphoxide (i) A solution of .. 1 »g. 0902241101) .._p-n-Phigrobenzyl-hetawctlomethyl-Zalia-metgglf-6-phenyl-bromine The methylene chloride solution was stirred with 473 mg (0.0022 mol) of m-chloroperoxybenzoic acid. The solution was stirred for 3 hours at 230 DEG. The methyl chloride was evaporated to 20 ml at 15 mm and 33 ° and the concentrated solution was diluted with 50 m1 "Ske11yso1ve B".

The solvent was decanted and the residue was taken up in ether, the product 4 soon crystallized in a yield of 250 mg (24%), m.p. 136-1370 (dec.).

Ana1ys Ber. for C 15 H 14 BrCl 1 N 2 O 6 S: C 38.68 H 3.02 N 6.02 Found: C 39.14 H 3.13 N 5.96 IR (KBr): 1800 (s), 1760 (s), 1520 (s) , 1350 (s), 1200 (s), 1050 (m), 830 (w), 740 (w) cmJ. 1 H-NMR (60 mHz, DMSO): δ 1.32 (s, 3H), 3-8-4.5 (m, 2H), 4.97 (s, 1H), 5.25 (d, J ~ 1.5 Hz, 1H), 5.45 (s, 2H), 5.6 (d, J ~ 1.5 Hz, 1H), 7.8-8.5 (m, 4H). _. f. IU 466 203 Potassium-Zbeta-clonethyl-Zalfa-methylpenam-3alpha-carboxylate sulfone (§) BL-Pzors) To a solution of 7 g (0,055 mol) of p-nitrobenzyl-2beta-chloromethyl-2alpha- -methyl- Alpha-bromopenam-3alpha-carboxylate sulfoxide (4) in 150 ml of ethyl acetate was added to a suspension of 4 g of 30% palladium on diatomaceous earth ("Celite") and 2.8 g of sodium bicarbonate in 150 ml of water. The mixture was hydrogenated for 3 hours at a pressure of 3.5 kg / cm 2. The catalyst was removed by filtration and the aqueous layer was separated and treated with 1.5 g of potassium permanganate in 50 ml of water. The mixture was stirred for 1 hour and 250 mg of sodium bisulfite were added. The mixture was filtered and the filtrate was adjusted to pH 2 with concentrated hydrochloric acid. The solution was lyophilized to give a white amorphous powder. The solid was extracted with ethyl acetate, evaporated to a volume of 20 ml and diluted with 100 ml of Skellysolve B. White, hygroscopic solid 2beta-chloromethyl-2alpha-methyl-penam-3alpha-carboxylic acid sulfone was recovered. The acid was dissolved in acetone and treated with solid potassium 2-ethylhexanoate. A crystalline white salt precipitated and, after filtration, 170 mg of the product were obtained with a melting point of aversr1ganae «14o9» çsanaerde1n1ng;

Anal vs Ber. for C 8 H 7 ClKNO 5 S · 2H 2 O: C 28.27 H 3.24 N 4.12 Found: C 28.27 H 3.69 N 3.84 IR (KBr): 1790 (s), 1770 (m), 1620 (s) 1460 (m), 1370 (s), 1310 (s), 1200 (s), 140 (s), 955 (m), 740 (m) cm -1], 1 H-NMR (10 0 mHz, D 2 O): δ 1.68 (s, 3H), 3.2-3.9 (m, J ~ 2 Hz, J ~ 4 Hz, J ~ 6 Hz, 2H), 4.0-4.4 (m, 2H ), 4.3 (s, 1H), 5.02 (dd, J ~ 4 Hz), JN2 Hz, 1H).

Example 2 Pivaloyloxymethyl-2beta-chloromethyl-2alpha-methylpenam-3alpha-carboxylate sulfone 2beta-chloromethyl-2alpha-methylpenam-3alpha-carboxylic acid sulfone in dimethylformamide was treated with one equivalent of triethylamine and stirred to give solution. dimethylformamide was then added. The resulting mixture was stirred at room temperature.

The mixture was then clarified by filtration and the filtrate was poured into ice water. The separated solid was collected by filtration, washed with water and dried to give the title ester.

The respective acetoxymethyl, methoxymethyl, acetonyl and phenacyl esters of true acid were prepared by replacing the bromomethyl pivalate used therein with an equimolar amount of chloromethyl acetate, chloromethyl methyl ether, chloroacetone and phenacyl bromide, respectively.

Example 3 Pivaloyloxymethyl-Zbeta-chloromethyl-Zalfa-methylpenam-3alpha-carboxylate sulfone (L1-iß) QO \\ s // C33 \\\\\ o CH C1 '_ "_ acetone (306) -_ 0 CO2 ° K ( BL ~ P20l3) 1 Q \ 0 CH _12 $ ”\ ~ ° 3 låF ___ 42. CHZCI O l 0 cozcaz-o-å-c (css) 3 (BL-P2024).

To a stirred suspension of 14.6 g (0.0487 mol) of the compound BL-P2013 (§) in 200 ml of acetone was added 4 ml of a 10% aqueous solution of sodium iodide and the mixture was refluxed on a steam bath. To the refluxing suspension was added 14.8 ml (0.1 mol) of redistilled chloromethyl pivalate (b.p. 34 ° at 7 mm Hg) in one portion. The mixture was stirred at reflux for 3 hours and then cooled to room temperature (22 °). The crystalline solid was collected by filtration and washed with 3 x 30 ml of acetone and the combined filtrates were evaporated to an oil in vacuo at a temperature below 22 °. The oil was then taken up in 500 ml of ethyl acetate and washed once with 200 ml of water and once with saturated sodium sulfate solution with stirring with 2 g of decolorizing carbon and under cooling (ice bath). After 20 minutes, the mixture was suction filtered through a diatomaceous earth pad ("Dicalite") and the pad was washed with 4 x 100 ml of ethyl acetate. The combined filtrates were concentrated in vacuo at 22 ° to an oil. The oil was then further concentrated at about 22 ° and at a pressure below 1 mm Hg to remove most of the remaining chloromethyl pivalate. The residual oil was then triturated twice with 50 ml portions of n-pentane and then allowed to stand over the weekend in a cold room (about 1 ° °) under n-pentane. The solid crystalline mass was then decomposed into a solid powder under 40 ml of a 4: 1 mixture of ether-n-pentane. The product was collected by filtration, washed with a 1: 1 mixture of ether and pentane and then with pentane and air dried. After drying in vacuo for 5 hours over P2 O5, 1.3.37 g of pivaloyloxymethyl-Zbeta-chloromethyl-2alpha-methylpenam-3alpha-carboxylate sulfone were obtained (about 75% yield) with a melting point of 93-95 °.

Analysis Ber. for C 14 H 20 ClNO 7 S: C 44.03 H 5.27 N 3.67 Found C 44.11 H 5.08, N_3.85. Example 4 Recrystallization of potassium 2-beta-chloromethyl-Zalfa-methylpenam-3alpha-carboxylate-sulfone (BL-P2013) To a mixture of 20 ml of n-butanol and 1g of the compound BL-P2013 (§) was added water, 1 ml at a time, from a separatory funnel with shaking until a pale yellow solution was obtained. The clear solution was filtered through a pleated filter paper and the flask and filter paper were washed with approx. 10 ml of a 9: 1 mixture of n-butanol and water and the combined filtrates were diluted with a further 20 ml of n-butanol. The resulting solution was introduced into a round bottom flask in a rotary extender and evaporated under reduced pressure to about half the original volume. The snow-white crystalline product was collected by filtration, washed with 6 x 10 ml of acetone and air-dried. Yield 810 mg. After vacuum drying for 6 hours over P2 O5 at a pressure of less than 1 mm Hg, 800 mg of product with a melting point of 155 ° (decomposition) were obtained, which corresponded to a yield of 80%.

Anaïvs Ber. for C 8 H 9 ClNO 5 SK-1H 2 O: C 29.67 H 3.39 I N 4.63 C1 10.94; K.F.H20, 5.56.

Found: C 29.23 H 3.38 N 4.49 Cl 10.74; K.F.H20, 5.74.

This recrystallization procedure gave a crystalline monohydrate which differs from the starting material, which was substantially anhydrous.

Exemgeï 5 3 'å cH3o-c-c1 f312 E clcaê-o-c-cl [jag + c1so H i> 3 __ 6: a o, se chemim Abstract; _21, 2427 'och _22, 3828 sam es 299064. å * Öí-'Hw-NH g3C "“ c, fNÉ ~. _ L! EK \\ cfm; | \ o OO! Um m 466 203 6 clsozcngcl (_) NaI acetone Që 49 HÉCI ff 0 g-øcag: Ou- [03 SC II // lc l / l, COOK (ILS. 3: 3l6,2ll7) 466 203 _ 22 c cH-co_.m; S H3 I __ \ l "" N "ß Ha 0033 o 22 o -o §s4 cngcl I 1,033 IC o / 'J -' @ ..____ 0 Il o I HI“ s 03-- .__ S * CO NH, ..__ T4 ”oc NI ¿... “ß * N32 'o °"' co. * Ï 'o AE ° § 1/0 c c1 * s H2 - 1 kan C open-N x, 3 I Il 23 466 203 A solution of 0.115 mol of chloromethyl chlorosulfate in 40 ml of dichloromethane was added dropwise, while maintaining the reaction temperature below 30 °, to a solution of 0.1 mol of compound 5, 0.3 mol of potassium bicarbonate and 0.01 mol of tetrabutylammonium hydrogen sulphate in 200 ml of a 1: 1- At the end of the addition, the mixture was stirred at room temperature for 30 minutes, the organic phase was separated and the aqueous phase was extracted with 50 ml of dichloromethane. The combined organic phases were dried over sodium sulphate and evaporated in vacuo to give a mixture of dichloromethane. of a residue, which was dissolved in 150 ml of ether. Insoluble material was filtered off after addition of diatomaceous earth and the filtrate was evaporated in vacuo to give compound 7. b) To a suspension of 1.5 g of compound 5 in 12 ml of dimethylformamide was added 1.6 g of bischloromethyl sulfate and the mixture was stirred at room temperature for 45 minutes. After dilution with 50 ml of ethyl acetate, the mixture was washed with water and then with an aqueous solution of sodium bicarbonate, dried and evaporated in vacuo to give compound 7 as an oil. c) To a solution of 0.005 mol of compound 5 in 7.5 nl of dinethylformamide was added 0.007 mol of triethylamine and 0.030 mol of chloroiodomethane and the ignition was stirred for 4 hours at room temperature. After dilution with 30 ml of ethyl acetate, the mixture was washed with 3 x 10 ml of water, followed by 5 ml of a saturated aqueous solution of sodium chloride, dried and evaporated in vacuo to give compound 7 as an oil. d) To a mixture of 0.15 mol of compound 5, 0.15 mol of silver nitrate and 7.5 g of silver oxide in 750 ml of acetonitrile was added 1.5 mol of chloroiodomethane. After stirring for 48 hours at room temperature, the silver salts were filtered off and the filtrate was evaporated to dryness in vacuo. The residue was dissolved in 200 ml of ethyl acetate and the solution was washed with a saturated aqueous solution of sodium chloride, filtered, dried and evaporated in vacuo to give compound 7.

Compound 7 and other intermediates and final products of the present invention are purified, if desired, by column chromatography, such as on Sephadex LH 20 using a 65:35 mixture of chloroform and 466,205 M hexane as eluent, or by silica gel chromatography, for example using Maillin-rooted CC-7 and a 3: 2 mixture of hexane and ethyl acetate, an 8: 2-, 7: 3-, 1: 9 or 15:85 mixture of ethyl acetate and petroleum ether, a 4: 6 or a 3: 1 mixture of ethyl acetate and n-hexane, a 3: 1-, 1: 1 or 1: 4 mixture of hexane and ethyl acetate or a 1: 1 mixture of cyclohexane and ethyl acetate.

Thin layer chromatography is also useful. "Sephadex" is crosslinked dextran-2- (diethylamino) -ethyl-2- [Z2- (diethylamino) ethyl diethylamine] ethyl ether hydrochloride hydrochloride piclorohydrin (see Merck Index, 9th Edition, No. 7337).

A solution of 0.2 mol of compound 7 and 0.3 mol of sodium iodide in 150 ml of acetone was stirred for 18 hours at room temperature. The resulting suspension was cooled to about 0 ° and adjusted to pH 7.2 by the addition of a saturated aqueous solution of sodium bicarbonate with stirring. After decolorization by titration with 0.5 M of an aqueous solution of sodium thiosulfate, 150 ml of water were added dropwise to the stirred mixture to give the solid compound 8, which was taken up by filtration, washed with 2 x 20 ml of a 1: 1 mixture. of acetone and water.a2axaZ fl mml.isophi_-propanol1 and 2 X 20 "m1 ether as well as dried sieve Ampici11in was converted to compound 9 by using methyl acetoacetate in the process according to U.S. Patent 3,316,247. Then a stirred solution of 0.57 was added to a stirred solution. of compound 9 in 1 liter of dimethylformamide 0.5 mol of compound 8 at 5 ° After stirring for 15 minutes at 5 °, the reaction mixture was poured onto an ice-cold mixture of 4 liters of ethyl acetate and 2 liters of a saturated aqueous solution of calcium chloride with stirring. The layer was separated, washed with 2 x 500 ml of a saturated aqueous solution of calcium chloride, filtered and evaporated to about 1 liter in vacuo to give a concentrated solution of compound 10. To this concentrate was added 500 ml of water and 500 ml of n-butanol, and then 4 N hydrochloric acid was added dropwise with stirring until the amino-protecting group had been removed to obtain a solution of compound 11. After complete addition of the acid, 1 ml was added. 1 liter of ether and 500 ml of water for the stirred mixture, the aqueous phase was separated and the organic phase was extracted with 800 ml of water. The combined aqueous extracts were washed with 1 liter of ether and then 640 g of sodium chloride and 2 liters of dichloromethane were added and the mixture was stirred for 15 minutes. The organic phase was separated and the aqueous phase was extracted with 1 liter of dichloromethane and the combined organic extracts were dried over magnesium sulphate and evaporated to about 600 ml under reduced pressure to obtain a concentrated solution of compound II. Addition to the concentrate of 200 ml of 2-butanone, followed by cooling, precipitated solid 6-LIR) -2-amino-2-phenylacetamide] -3,3-dimethyl-7-oxo-4-thia-1-aza- Bicyclol.2.2Hheptane-2-carbonyloxymethyl-2beta-chloromethyl-2alpha-methylpenam-3alpha-carboxylate sulfone (II), which was recovered by filtration.

Example 6 6-E1R) -2-amino-2-p-hydroxyphenylacetamide] -3,3-dimethyl-7-oxo-4-thia-1-aza-bicyclo [3.2.1] heptane-2-carbonyloxymethyl-Zbeta-chloromethyl -2-alpha-methylpenam-3-alpha-carboxylate sulfone of the formula Ho-Ö- cH-co -NH ._ 1 A 4 _NH2 ° ___u o * o. Was prepared by replacing ampicillin with amoxicillin in the procedure of Example 5. 466 zos - N Exemgeï 7 a> i * a qi Ü * N-bromosuccinimide \ O a-azb-isobutyronitrile CClu C * ia C / E. \ \ o In accordance with U.S. Patent 3,860,579, 50 g (0.375) were refluxed. moi) recrystallized phthalide and 0.375 moi recrystallized N '-bromsuccirnimidf4, .5'.atimer: ï <-i: f.- nämamræv, æaaæmik =: nagï,; - ïæIfa-azobutyronrïatri'Ia 1' i 1 iiter cci4. The mixture kyides: iii about ice 'acñ was filtered to remove succinimide, which was washed with about 100 ml of CCl 4 and filtered. The combined CCl 4 phases were concentrated in vacuo to about 150 ml to give solid 3-bromophthalide, which was taken up by filtration, washed with about 50 ml of CO 14 and dried to give 54 g, the product weight being reduced to 50 g after recrystallization from cocaine. - the cycionexan. melting point 84-as °.

CI fl Br o 'o - \! ca c1 \ 2 _ o I ”a 'l + -r i: H3 a: 12 ær-I-N _ t,. o "_" a c-ox - u ä. °.

INEF 22 '2; 466 203 o '/ o \ s / cnzcl - ”m3 .H -. To a stirred partial solution and partial suspension of 2.3 g (0.0075 moD of compound 5 (BL-P2013) in 20 ml of dimethylformamide (DMF; dried at least 3 weeks over 3A molecular sieve) 1.7 g (0.008 mol) of 3-bromophthalide (12) were added and the mixture was stirred for 4 hours at 220. The resulting mixture was poured into a mixture of 200 ml of ice-cold water and 200 ml of ice-cold ethyl acetate (flask rinsed with a small amount of ethyl acetate) and the mixture was shaken.

Then the organic solvent phase was separated and washed with 7 portions of icy water (JOD ml). The ethyl acetate phase was washed; a saturated aqueous solution of sodium sulphate, dried in cold over sodium sulphate, filtered and evaporated to dryness in vacuo to give the residue an oil which was triturated twice with 25 ml of methylcyclohexane, twice with 25 ml of "Skellysolve B" and four times with 25 ml of n-hexane to give 2.5 g of compound 13 as an almost white solid after drying in air, this product was dried over P 2 O 5 at a pressure of less than 1 mm Hg to give 2.5 g g of compound 13 with a melting point of 104 ° (decomposition), its estimated purity was 85-95 °.

Analysis Ber. for C 16 H 14 ClNO 7 S: C 51.61 H 3.79 N 3.77 Cl 9.53 Found: C 52.59 H 4.67 N 3.21 Cl 7.73; K.F.H2U, 0.27. 466 203 æ Example 8 Pivaloyloxymethyl 2-beta-chloromethyl-2alpha-methylpenam-3alpha-carboxylate sulfone A mixture of 1g (0.003l mol) of potassium 2-beta-chloromethyl-2alpha-methylpenam-3-alpha-carboxylate sulfonyl hydride and 1g in 15 ml of dimethylacetamide for 2 hours at 23 °. To this mixture was added 470 mg (0.003l mol) of pivaloyloxymethyl chloride and stirring was continued for 18 hours.

The molecular sieve was collected and the filtrate was diluted with 100 ml of water and extracted with ethyl acetate. The ethyl acetate was washed nine times with water and dried over anhydrous magnesium sulfate. The solvent was removed at 30 ° (15 mm Hg) to give an oil which was chromatographed on silica gel using Silicar CC-7 (8 parts methylene chloride, 2 parts ethyl acetate) to give a spot with an Rf value of 0.5. The resulting residue was crystallized from heptane ("Skellysolve B") to give 100 mg of pivaloyloxymethyl-2beta-chloromethyl-2alpha-methylpenam-3alpha-carboxylate sulfone, m.p. 94-950. É fl šllší ~ ~ Ber. : C 44.20 H 5.4 N 3.63 NMR and IR spectra were consistent with structure.

Example 9 Sodium 2-beta-chloromethyl-2-alpha-methylpenam-3-alpha-carboxylate-sulfone Qšyå gg cá.cl H + Nggg 'q§ {íÉQ'CH2Cl Ff ”en _ (2 _) _> III [C33 3 - N Å * N y To a stirred solution of 500 mg of the compound BL-P 2 O 3 (the potassium salt) N 466 203 I in 5 ml of water and 10 ml of ethyl acetate was added 2N HCl until a solution of 1 .mu.l was obtained. 2K c8H9c1No5sNa (396) - <289.6v pH value of 1 (performed in ice bath with vigorous stirring). The mixture was then saturated with sodium sulfate, the aqueous layer was separated and the organic phase was rapidly dried in ice over sodium sulfate, filtered and treated dropwise with 50% NaEH (sodium 2-ethylhexanoate) in anhydrous n-butanol until a neutral reaction could be detected with moistened pH paper.

The product did not crystallize on scraping and on concentration of the mixture in vacuo an oil was obtained which was dissolved in 5 ml of acetone. Upon scraping, no crystals were still obtained, nor when ether was added to the cloud point. Concentration in vacuo on a rotary evaporator to an oil dissolved in ethyl acetate also did not yield any crystals on scraping after the addition of a drop of water. Concentration in vacuo and trituration of the residue with 5 ml of n-butanol gave 200 mg of an amorphous white powder, which was washed with ether, air dried and vacuum dried over P 2 O 5 for 24 hours. This gave 180 mg of sodium 2-beta-chloromethyl-2alpha-methylpenam-3alpha-carboylate sulfone with an indeterminate decomposition point exceeding 100 °.

Analysis; _ V Ber; for C8H9CTN05SNa: 'C 33, 10 "ß H SJT3' N 4; 89" »¥ Found: C 33,20 H 3.69 N 4.44 KFH2O, 4.04 Example 10 Potassium-Zbeta-chloromethyl-2- Methylpenam-3-carboxylate sulfone (BL-P2013) To 10 liters of water, 130 g (1.25 moles) of sodium bicarbonate and 200 g of 0% Pd on barium sulfate was added 272 g (0.565 moles) of p-nitrobenzyl-6alpha-bromo- -Zbeta-chloromethyl-2-methylpenam-3-carboxylate sulfone, dissolved in 5 liters of ethyl acetate. The mixture was hydrogenated at 400 and a pressure of 1 kg. After hours, hydrogen uptake became very slow and 200 g of 10% Pd on barium sulphate was added, after which the mixture was hydrogenated until no further significant hydrogen absorption could be detected.

The slurry was filtered through a pad of diatomaceous earth (Celite) and the pad was washed with water, after which the aqueous phase was washed with 3 liters of ethyl acetate. To the aqueous solution was added 3 liters of ethyl acetate and the pH of the mixture was charged at 1.5 with iso ml of 12 N Hci at 10 °. The organic phase was separated and the aqueous solution was saturated with Na 2 SO 4 · 10 H 2 O and extracted with 2 x 1 liter of ethyl acetate. The combined extracts were dried over methyl chloride. The drying medium was removed and 260 ml of 2 N potassium 2-ethylhexanoic acid in butanol was added at 0 DEG.

After stirring for 2 hours at 0 ° C, potassium 2-beta-chloromethyl-2-methylpenam-3-carboxylate sulfone (BL-P2013) was taken up and dried in vacuo at room temperature. Yield 134.8 g (ca. 70%).

Exemgei 11 p-nitrobenzyl-6a1fa-bromopenicylamate-sulfoxide o -lär gg ”ff l /. + TEA + 131-032 -G-Noz (101) -n-w I / -iiw _. (295) eL; TßS: f Ä "'N eo cozcnz-Q NO2; n31¿2s) Ti11 200 ml of N, N-dimethylacetamide was added 44 g (0.148 mol) of 6a1fa-bromopenicyl-nanoic acid sulfoxide, followed by 20.5 (0.148 mol) of triethylamine and 38.2 g (0.177 mol) of p-nitrobenzyl bromide The mixture was stirred for 20 hours at 22 DEG C. The reaction mixture was poured into 1 liter of water and extracted with 3 x 300 ml of methylene chloride. was taken with 200 ml of a 5% aqueous solution of sodium bicarbonate and dried over sodium sulphate for 1 hour at 5 °. The solution was filtered and evaporated in vacuo to a residue. The residue was diluted with ether and the solid was collected by filtration. 54 g of p-nitrobenzyl-6-alpha-bromopenicillanate sulphoxide were obtained after drying in a yield of 85% NMR was consistent with the structure.

The yield in this step was the same as in the esterification of the K-salt. The advantage of this step is that there is no need to produce any K-salt (the yield in this step is 85-90%).

Example 12 p-Nitrobenzyl 6-alpha-bromopenicillanate sulfoxide To 4.375 liters of N, N-dimethylacetamide was added 873.0 g (2.95 mol) of 6-alpha-bromopenicillanic acid (S) -sulfoxide and then added, with a mixture of fl9.0 C H while maintaining the internal temperature below 35 °; 293 g (2} 95 mol) of tri% '' ethylamine followed by 764 g (3.54 mol) of p-nitrobenzyl bromide. The mixture was then stirred at room temperature for 5 hours and allowed to stand overnight.

The reaction mixture was poured into 20 liters of water and extracted with 3 x 7 liters of methylene chloride. The combined organic extracts were washed with x 7 liters of water and then with 7 liters of a 5% aqueous solution of sodium bicarbonate and dried over anhydrous magnesium sulfate. The magnesium sulfate was filtered off and the solution was evaporated to a crystalline residue; 4 liters of diethyl ether were added and the crystals were collected to give, after drying at room temperature, 177 g (92%) of p-nitrobenzyl-Galpha-bromopenicillanate sulfoxide.

Found: Br 188.48% (calc. 18.53%); ° (D (0.25% MeOH) + 1620. 466 203 32 Example 13 N-nitrobenzyl-alpha-bromo-Zbeta-chloromethyl-2-methylpenam-3-carboxylate sulfone To 16 liters of acetic acid was added 364.6 g (0 812 mol) of p-nitrobenzyl-6-alpha-bromo-Zbeta-chloromethyl-2-methylpenam-3-carboxylate To the solution thus obtained and stirred at room temperature was added dropwise over 3 hours a solution of 282 g (l , 78 mol) of potassium permanganate in 26 liters of water.

The mixture was then stirred at room temperature for 1 hour and 37% hydrogen peroxide was added dropwise until a colorless solution was obtained. liters of water were then added, the mixture was stirred for 1 hour at room temperature and the crystalline precipitate was collected, washed with 3 x liters of water and with 2 x 2 liters of ethanol and dried in vacuo at room temperature. Yield 297 g (76%). an (msz cnzciz) + 7s, 9 ° Example l4 St-P20l3 as free acid; §gfj c cl 11201 ”à H2 H-fpoll- 'IC. - n »1 H3 -> H; v- f, e e - ,, f! To a mixture of 25 ml of ethyl acetate and 10 ml of water was added 800 mg (0.00261 mol) of the compound BL-P2013 as the potassium salt. After all solids had dissolved, the mixture was treated dropwise with a 50% aqueous solution of phosphoric acid with vigorous shaking until no further material precipitated from the aqueous layer. The ethyl acetate layer was separated, then washed with a saturated sodium chloride solution and dried over anhydrous magnesium sulfate. The desiccant was removed by filtration and washed with 10 ml of ethyl acetate. The washing solvent was combined with the original filtrate. "Skellysolve B" was then added to the ethyl acetate until the cloud point was reached (about 10 ml). The mixture was treated with 500 mg of activated carbon ("Darko KB") and filtered. The filtrate was diluted with 15 ml of "Skellysolve B" and then added with seed crystals of the compound BL-P2013 in the form of the free acid. After approx. For 3 hours at room temperature, the crystalline precipitate of the free acid was collected and dried in vacuo (15 minutes) over P 2 O 5 to give 323 mg (46%). The product melted during slow decomposition over 10 ° C.

Analysis Ber. for C 8 H 10 ClNO 5 S: C 35.89 H 3.77 N 5.23 Cl 13.25 Found: C 35.88 H 3.91 N 5.41 Cl 13.52 The product was found to be unstable when stored at 23 ° for 7 hours. day.

Example 15 oalpha-bromopenicillanic acid sulfoxide fl, f 'BI-lea s _ Bra' S (G6 CHZNHC) 2, __, _ \ @___ .4CO HÉ Hé> N. a- '. MQOQII W8 °° ß4 MV 296.14 To 3 liters of methylene chloride was added 300 g (0.75 mol) of 6alpha-bromopenicillanic acid N, N'-dibenzylethylenediamine salt and this suspension was cooled to 5 °. Then 130 ml of concentrated hydrochloric acid were added dropwise over 15 minutes and with good stirring. The mixture was then filtered through a pad of diatomaceous earth ("Celite") and the filter cake was washed with 3 x 250 ml of methylene chloride. The combined methylene chloride solutions were washed with water (2 x 500 ml) and dried over sodium sulphate for 15 minutes. The 466,205 M sodium sulfate was removed by filtration and the filtrate was evaporated under reduced pressure to a volume of about 750 ml.

The resulting solution was cooled to 50 and with vigorous stirring, 130 ml of 40% peracetic acid was added dropwise in such a way that the temperature was kept at 5-122 °. The reaction was strongly exothermic. Towards the end of the addition, the slurry was stirred for 2 hours at 5 ° and the product was collected by filtration and washed with 100 ml of cold water (50) and 100 ml of cold methylene chloride (5 °). 166 g (57%) of 6-alpha-bromopenicillanic acid sulphoxide, m.p. IR and NMR spectra were consistent with the structure of the desired product.

Analysis Ber. for C 8 H 10 BrNO 4 S: C 32.44 H 3.40 N 4.73 Found: C 32.30 H 3.35 N 4.7l H 2 O, 2.18 Potassium 6-alpha-bromopenicillanate sulfoxide. a -. '' __ __¿. ~ V, f? ', -. i, _ _ Bål-y; S _r; r - i r- ”0023. wc 021: Mv: zman To 3 liters of acetone were added 26 g (0.43 mol) of 6alpha-bromopenicillanic acid sulfoxide and 162 ml of 50% (wt%) potassium-2-ethylhexanoic acid in n-butanol. After stirring for 1 hour at 220 DEG C., the product was collected by filtration, washed with 2 x 250 ml of acetone and dried. 277 g (90%) of potassium galpha-bromopenicillanate sulfoxide with a melting point of 185 DEG C. were obtained. IR and NMR spectra were consistent with the desired structure.

Analvs Ber. for C 8 H 9 BrKNO 4 S: C 28.75 H 2.71 N 4.19 Found: C 29.03 H 2.78 N 4.04. ß 466 203 ggnitrobenzyl-6alpha-bromopenicillanate-sulfoxide Br § Br 2 wo '+ Il' ÛICQ K v-- o Q 2 y cozc 02 uv 431.28 To 1 liter of N, N-dimethylacetamide was added l45 g (0.43 mol potassium 6-alpha-bromopenicillanate sulfoxide and with stirring then 115 g (0.53 mol) of p-nitrobenzyl bromide were added at 22 °. The mixture was stirred for 20 hours at 22 °.

The reaction mixture was poured into 3 liters of water and extracted with 3 x 1500 ml of ethyl acetate. The combined ethyl acetate extracts were washed with 2 x 500 ml of a 5% aqueous solution of sodium bicarbonate and dried over sodium sulfate, for 0.5 hours; The sodium sulfate was filtered off and the filtrate was evaporated under reduced pressure to a residue, to which 1 liter of diethyl ether was added, which caused the product to crystallize. The crystals were collected by filtration, washed with 2 x 100 ml of diethyl ether and dried to give 162 g ( 87%) p-nitrobenzyl 6-alpha-bromopenicillanate sulfoxide, m.p. 11 ° C. IR and NMR spectra were consistent with the desired structure.

Analvs Ber. for C 15 H 16 BrN 2 O 6 S: C 41.78 H 3.5l N 6.50 Found: C 411.66 H 3.45 N 6.85 H 2 O, 0.69. p-Nitrobenzyl 6-alpha-bromo-2beta-chloromethyl-2-methylpenam-3-carboxylate O - '. _ Br š 'Br' _ lz \ 4 "* c F - IW 466 203% To 1 liter of p-dioxane was added 70 g (0.66 mol) of p-nitrobenzyl-6alpha-bromopenicillanate sulfoxide, followed by 2l , 2 ml (0.1 mol) of benzoyl chloride and 21.8 ml (0.1 mol) of quinoline. The reaction mixture was refluxed for 4 hours and then cooled to 22 °, poured into 2500 ml of water and extracted with 3 x 800 ml of ethyl acetate. The combined ethyl acetate extracts were washed with 300 ml of a 5% aqueous solution of sodium bicarbonate, 300 ml of a 5% aqueous solution of phosphoric acid and 300 ml of water. The ethyl acetate solution was dried over sodium sulfate for 0.5 hours and the sodium sulfate was removed by filtration. The filtrate was evaporated under reduced pressure to a residue, which was redissolved in 1 liter of ethyl acetate and again evaporated under reduced pressure to a residue. Then 1 liter of diethyl ether was added and the product was collected by filtration in a yield of 41 g (57%) of p-nitrobenzyl-6alpha-bromo-2beta-chloromethyl-2-methylpenam-3-carboxylate, m.p. IR and NMR spectra were consistent with the desired structure. 0 Analysis Ber. for C 15 H 14 BrClN 2 O 5 S: C 40.06 H 3, 14 N 6.23 Found: - - C 1 - 4. 11 - 11. * p-nitrobenzyl-6-alpha-bromo-Zbeta-chloromethyl-2-methylpenam-3-carboxylate sulfoxide 0 i c 31,; s 3201 Bry 'S H2 ° / I I' II ”of '/ Éfo c - Ü. f? To NO200 ml of methylene chloride was added 51 g (0.1 mol) of p-nitrobenzyl-6alpha-bromo-2-beta-chloromethyl-2-methylpenam-3-carboxylate, followed by 23 g (0.2 mol) m-chloroperoxybenzoic acid. The solution was stirred for 2 hours at 22 ° and evaporated under reduced pressure to a moist residue. The residue was stirred with 4 liters of diethyl ether for 1 hour and allowed to stand for 20 barrels at 10 °. The product N 466 205 crystallized out and was collected by filtration, washed with 2 x 200 ml of diethyl ether and dried to give 39 g of p-nitrobenzyl-Galfa-bromo-2beta-chloromethyl-2-methylpenam-3-carboxylate sulfoxide (75%) with melting point 322 °.

IR and NMR spectra were consistent with the desired structure.

Anal vs Ber. for C 15 H 14 BrClN 2 O 6 S: C 38.69 H 3.03 N 6.07 Found: C 38.98 H 3.04 N 5.84 H 2 O, 0.35.

Potassium Zbeta-chloromethyl-2-methylpenam-3-carboxylate sulfone (BL-P2013) O.

Bg, š C fi zßl _ x / f-'Hzf-'l “L Tee“ t -fi I 'm5 / 1 / __ 0 ,, <5 / ° I ° 2 CRTQ- N02 J * f-cozx To 600 ml water 8 g of 30% Pd was added to Celite and 16 g (0.6 mol) of sodium bicarbonate, then 32 g (0.69 mol) of p-nitrobenzyl-6-alpha-bromo-2-beta-chloromethyl-2-methylpenamide were dissolved. 3-Carboxylate sulfoxide in 400 ml of ethyl acetate and added to the aqueous slurry. The mixture was hydrogenated in a Parr apparatus at a pressure of 3.5 kg / cm 2 at 22 ° for 4 hours. The slurry was filtered through a thin pad. Celite "pad on a sintered glass funnel, the pad was washed with 2 x 50 ml of water and the aqueous layer was separated from the combination of filtrate and washings. The aqueous layer A was washed with 200 ml of diethyl ether and then cooled to 5 ° and with stirring a solution of 12 g (0.076 mol) of potassium permanganate in 200 ml of water for 0.5 hours, keeping the pH between 7.5 and 8.0 by adding 40% phosphoric acid, when the light red color did not disappear after 5 minutes was not added n eyes more potassium permanganate solution. The reaction mixture was stirred with a small amount (about 50 mg) of sodium bisulfite for 0.5 hours and then the slurry was filtered through a pad of 466,205 W of a "Celite" pad. The pad was washed with 2 x 50 ml of water. The combination of filtrate and washings was coated. with 500 ml of ethyl acetate and with stirring the pH was adjusted to 1.5 by adding 2 N hydrochloric acid, the layers were separated and the aqueous layer was saturated with sodium sulfate, it was re-extracted with 2 x 400 ml of ethyl acetate and the combined ethyl acetate extracts were dried over sodium sulfate. 5 ° for 0.5 hours, the sodium sulfate was removed by filtration and the filtrate was evaporated under reduced pressure to a residue, the residue was dissolved in 160 ml of acetone and 160 ml of diethyl ether and 50% by weight of potassium 2-ethylhexanoate in n-butanol was added until The potassium salt of the compound BL-P2013 was crystallized out, filtered by filtration, washed with diethyl ether and the solution showed a neutral reaction on testing with moist pH paper. dried. The yield was 16 g of potassium Zbeta-chloromethyl-2-methylpenam-3-carboxylate-sulfon (BL-P2013) (76%) with melting point 2020. IR and NMR spectra were consistent with the desired structure.

Analysis Ber. for C 8 H 9 ClKNO 5 S: C 31.42 H 2.97 N 4.58.18; t _ .2ig.g_8§: 1'u = 40151 * - Q-ggt-atmvf; Example 16 Pivaloyloxymethyl-Zbeta-chloromethyl-2-methylpenam-3-carboxylate sulfone §BL-P2024! 'oo 0' 0 Rs / cazcl _ R ß H2 ° 1 o; - * ocozx ('- "' co'2o_1_1_2ococ (ci13) 3 BL-Paozu., lßjV 381183 __ _______ To a stirred suspension of 14.6 g ( 0.0487 mol) of potassium 2-beta-chloromethyl--2-methylpenam-3-carboxylate sulfone (BL-P2013) in 200 ml of acetone, 4 ml of a 10% aqueous solution of sodium iodide were added and the mixture was brought to reflux To the refluxing suspension was added 14.8 ml (0.1 mol) of redistilled chloromethyl pivalate (b.p. 34 ° at 7 .mu.m Hg) in one portion, the mixture was refluxed for 3 hours and then cooled to room temperature (220). the solid was collected by filtration and washed with 3 x 30 ml of acetone and the combined filtrates were evaporated to an oil under reduced pressure at a temperature below 22 ° The oil was then taken up in 500 ml of ethyl acetate and washed once with 200 ml of water and a batch with 200 ml of a saturated sodium sulphate solution, the solution was then dried rapidly over sodium sulphate under cooling (ice bath) and while stirring with 2 g of decolorizing carbon. After 20 minutes, the mixture was filtered through a pad of Celite and the pad was washed with 4 x 100 mL of ethyl acetate. The combined filtrates were concentrated under reduced pressure at 22 ° to an oil. The oil was then further concentrated at about 220 DEG C. and at a pressure below 1 mm Hg to remove the majority of the remaining chloromethyl pivalate. The residual oil was triturated twice with 50 ml portions of n-pentane and then allowed to stand over the weekend at about 10 ° below n-pentane. The resulting solid crystalline mass was then decomposed: fat powder = 40% nloav, enf4: 1-hlanduingmau, diethyl-WW ether and n-pentane. The product was collected by filtration; washed with a 1: 1 mixture of diethyl ether and n-pentane and air dried. After drying in high vacuum for 4 hours over P2 O5, 13.37 g of pivaloyloxymethyl-Zbeta-chloromethyl-2-methylpenam-3-carboxylate sulfone (BL-P2024) (75%) with a melting point of 93-950 were obtained.

Purification of BL-P2024 Approx. 3 g of crude BL-P2024 (obtained as described above) were dissolved in 5 ml of ethyl acetate, applied to a 4.5 x 40 cm column of silica gel (Mallinkrodt CC-7) and eluted with a 4: 1 by volume mixture of CH and ethyl acetate. The fractions showing a single spot at an Rf value of 0.84 (thin layer chromatography on silica gel plate with a 4: 1 mixture of CH 2 Cl 2 and ethyl acetate; 12 detection) were combined and concentrated under reduced pressure to 1.88 g of a crystalline solid material. A portion of this material (900 mg) was dissolved in 5 ml of ethyl acetate; the residual solution was filtered, diluted almost to the cloud point with petroleum ether ("Skellysolve B") and then stored at room temperature for 3 days. The formed crystals were collected by filtration, washed with petroleum ether and dried to give 560 mg. purified BL-P2024 with melting point 100-110 °.

Analysis Ber. for C 14 H 20 ClNO 7 S: C 44.03 H 5.27 N 3.67 Found: C 44.1 H 5.08 N 3.85 Example 17 The ammonium salt of BL-P 2 O 13 250 mg of the free acid of BL-P 2 O 13, dissolved in 20 ml of a 1: 1 by volume mixture of acetone and methanol, filtered to give a clear solution.

Anhydrous ammonium solution was prepared by adding 1 ml of ammonium hydroxide (30%, reagent grade) to 10 ml of a 1z volume mixture of acetone and methanol, and then 1 g of anhydrous magnesium sulfate was added to the solution with gentle stirring, and the mixture was filtered through a filter paper. To the filtrate of the free acid was gradually added about 2 ml of "anhydrous ammonium solution" and the whole was mixed well. 100 ml of diethyl ether were mixed in one portion with the resulting mixture to precipitate the ammonium salt of the BL- The white ammonium salt was isolated from the solvent and washed with 2 portions of 50 ml of diethyl ether each, The isolated powder was dried at 350 in a vacuum oven overnight.

Analysis data were as follows: Ber. % C 33.7 H 4.6 N 9.8 Found: C 33.66 H 4.63 N 1D, 12 The product was found to be dry when tested according to the Karl Fischer method.

Microscopic examination of the product showed that it was a crystalline substance.

Example 18 Non-hygroscopic sodium salt of BL-P2013 50 mg of the free acid of BL-P2013 was dissolved in 4 ml of a 1: 1 by volume mixture of acetone and methanol. The mixture was filtered to give a clear solution.

Sodium 2-ethylhexanoate solution was prepared by dissolving 40 mg of sodium 2-ethylhexanoate in 10 ml of a 1: 1 by volume mixture of acetone and methanol. To the filtrate of the free acid was added 10 ml of the sodium 2-ethylhexanoate solution and the whole was mixed well. 5 ml of diethyl ether were mixed in one portion with the resulting mixture to precipitate the sodium salt of BL-P2013. The white salt was slurried in diethyl ether for 1-2 hours and then separated from the solvent and washed with 3 portions of 5 ml of diethyl ether each. The isolated powder was dried at 300 in a vacuum oven overnight.

Example 19 Crystallization of the compound BL-P2013 V U -. \ = g // _ * omxrisf. 400 mg of the compound BL-P2013 was dissolved in a minimum amount of a 1: 1 by volume mixture of acetone and water and diluted with 10 ml of acetone, filtered, then diluted with acetone to about 25 ml and scraped and after 30 minutes the crystalline hydrate was collected by filtration, washed well with acetone, air dried and then vacuum dried at a pressure of less than 1 mm Hg overnight.

Anal vs Ber. for C 8 H 9 ClNOSK-H 2 O: C 29.67 H 3.39 N 4.63 Cl 10.94; H 2 O 5.55. Found: C 29.32 H 3.32 N 4.44 Cl 11.3; H 2 O 5.90. 466 205 f 42 Example 20 The N, N'-dibenzylethylenediamine salt of BL-P20133 BL-P2013 + 1/2 N, N'-dibenzylethylenediamine diacetate Recrystallized. in acetone-ether or CH 2 Cl 2:: CHB r-_] '1 ,, J co2x.c6H5cH2NH-c1O2 306 mg (0.00l mol) of the compound BL-P20l3 was dissolved in 7 ml water and was added to a solution of 110 mg (0.0005 mol) of N, N'-dibenzylethylenediamine diacetate 1 '7 ml of water. The mixture was stirred and the salt crystallized out and after stirring fi cæ. lüflåsminuter fi uppvaxatogs, salietsgenomsfifi-s, treringaoch ^ lofttorkädesffö% "erhällande fl avä3 fl0" mgmäv + N% N§ * ¿* bBH $ YTë fi ¥ $ ë fl å¶ * '* f aminsaltet av BL-P20l3' ca. and dilution with ether to the cloud point, 260 mg of air-dried and vacuum-dried material were obtained.

Analysis Ber. c 51.69 H 5.42 N 7.53 c1.9, ss runner: c 49.39 H 5.49 N 7.05 'c 8.96 H 2 O, 1.23 (KF).

Example 21 Chloromethyl ester of BL-P20l3 o o- \\ / 03201. + Clcxz-ofso cl 2 f- »CHB f_ (clgc fi acäcnahlvæ fl soug Q“ Icoox * 3 KH °° 3 605,7) å cnáclz 320 ü 466 203 0 AV o To a vigorously stirred mixture of 15.25 g (0.05 mol) of the compound BL-P2013 (5), 15 g (0.5 mol) KHCO 3 and 1.7 g (0.005 mol) of tetrabutylammonium hydrogen sulfate (Aldrich Chem. Co.) in a mixture of 50 ml of water and 50 ml of CH 2 Cl 2 were added dropwise a solution of 9.5 g (0.0575 mol) of ClCH 2 -O SO 2 Cl in 40 mL of CH 2 Cl 2. The temperature rose to 260 and after the addition (which took about 15 minutes) the mixture was stirred for another 30 minutes. As the product crystallized out, additional CH 2 Cl 2 (ca. 400 mL) was added to give a solution. The separated CH 2 Cl 2 layer and 50 ml of CH 2 Cl 2 wash were combined and dried over magnesium sulfate with stirring, after which 2 g of decolorizing carbon ("Darco KB") were added.

After about 30 minutes, the mixture was filtered and concentrated to about 50 ml and 150 ml of isoPf was added to the solution; then under reduced pressure. The resulting crystalline precipitate to A was taken up by filtration, washed well with isopropyl alcohol and air dried. After vacuum drying at a pressure of less than 1 mm Hg, 8.5 g of chloromethyl-2beta-chloromethyl-2-methylpenam-3-carboxylate sulfone (7) with a melting point of 116 ° (decomposition; darkens above 10 °) were obtained.

Anal ys Ber. for C 9 H 18 Cl 2 NO 5 S: C 34.18 H 3.5l N 4.43 Cl 22.43 Found: C 34.16 H 3.45 N 4.47 Cl 22.46; H 2 O, 0.33 (KF).

Estimated purity was 90-95%.

Iodomethyl ester of BL-P20l3 0 0 0 0 \\ ß CH Cl \\ // CH2Cl s 2 N I s a ». lll / CH; To a stirred mixture of 5 g (0,0l59 mol) of the chloromethyl ester of the compound BL-P20l3_ (7) in a stirred mixture. 25 ml of acetone were added with 3 g (0.02 ml) of sodium iodide. The resulting slurry was stirred for 17 hours and then cooled to about 0 °. 2 drops of a saturated aqueous solution of potassium bicarbonate were added and the mixture was diluted dropwise with water for 10 minutes until 50 ml had been added. The slurry underwent a sudden color change from yellow via gray and purple to black and therefore the crystals were recovered immediately by filtration and washed with a cold 1: 2 mixture of acetone and water, then with 3 x 10 ml isopropyl alcohol, diethyl ether and finally with n-pentane and air dried to give 5.55 g (91% yield) of the iodomethyl ester of BL-P 2 O 13 (8), m.p. 18-119 ° (dec.). Estimated purity was about 90%. 6-LKR) -2-amino-2-phenylacetamide] -3,3-dimethyl-7-oxo-4-thia-1-azabicyclo [2.2.1] heptane-2-carbonyloxymethyl-Zbeta-chloromethyl-2-alpha -methylpenam-3-alpha-carboxylate sulfone (II) (a Dane salt of ampicillin; see US-P 3,316,247) 45,466,203 δ-1H-co -NH 'f / N cc / \ ß 4 :, ocn; 0 _ o. §Sff 021201 I IIICH) ïH2 of '' '§-_0 o I s C33 C ca -co -qm __Tß (å, CH; NI 4- ,, N32' 0 _ IC = O 0 'E ° \ / Åo c1 To a cold ice-stirred stirred mixture of 5.46 g (0.01 mol) of the indicated Dane salt of ampicillin (9; var). solvated with one molecule of isopropyl alcohol) in 60 ml of acetone was added 4.08 g (0.01 mol) of the iodomethyl ester of BL-P 2 O 3 (8) and the resulting near clear solution was stirred for 5 hours with the ice bath removed after 30 minutes. The majority of the acetone was removed in vacuo on a rotary evaporator and the resulting concentrated solution was dissolved in 200 ml of cold ethyl acetate which was then washed with 2 x 50 ml of ice-cold water and 2 x 100 ml of a saturated aqueous solution of sodium sulphate. The ethyl acetate solution was then dried over sodium sulfate and filtered and the majority of the ethyl acetate was removed in vacuo on a rotary evaporator, the residue was triturated with 2 x 200 ml of dry diethyl ether and the resulting solid was collected. was taken by filtration to give 5.5 g of compound 10 as a light red powder.

This powder was stirred in a mixture of 50 ml of water, 50 ml of n-butanol and 20 ml of ethyl acetate while 6N hydrochloric acid was added dropwise to pH 2.5.

Then one or two drops of hydrochloric acid were added from time to time to maintain the pH at 2.2-2.5 for 45 minutes. When the pH no longer showed a tendency to rise, 100 ml of diethyl ether were added to the mixture with good stirring. The aqueous phase was separated and combined with, et al, other, 25 ml of aqueous 2-extract of the aqueous solution of the aqueous solution was filtered off with 50 ml of diethyl ether, which was then discarded.

The aqueous layer was then stirred vigorously under a layer of 100 ml of 2-butanone (methyl ethyl ketone) while sodium sulfate was added to saturate the aqueous layer. The 2-butanone layer was separated, dried over sodium sulfate for 30 minutes in an ice bath, filtered and concentrated in vacuo to near and dry. The residual oil was triturated to a solid with n-butanol, washed well with ether and then with n-pentane, air dried and vacuum dried over P 2 O 5 at a pressure below 1 mm Hg to give 1.6 g of 6- [1R) -2-Amino-2-phenylacetamide-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo [3.2.1] heptane-2-carbonylmethyl-2-beta-chloromethyl-2-alpha-methylpenam-3alpha-carboxylate -sulfone (11) in crude form. IR and NMR spectra were consistent with the structure of compound II but indicated that the product was not highly pure. This solid product was estimated to contain at least 40% and perhaps as much as 80% 6- [IR) -2-amino--2-phenylacetamide] 73,3-dimethyl-7-oxo-4-thia-1-azabicyclo [B]. 2.Q7heptane-2-carbonyloxymethyl-Zbeta-chloromethyl-2-alpha-methylpenam-3alpha-carboxylate -sulfone. vnn:; M 466 205 Example Gel 22 Improved Synthesis of BL-P2013 This process simplifies the production of BL-P2013 by eliminating the previous use of catalytic reduction.

Step l o Br, 1 Oz S I I '+' on cc13 + n1cyk1anexy1xarb ° ai1m1a. / - f, I 2 0 / 'cooH ° H Ä Pyriain cH2c12 m: (See p. 633 in Cephalosporins and Penicillins, edited by Edwin H.

Flynn, Academic Press, New York, l972). g (0.1 mol) of 6alpha-bromopenicillanic acid sulfoxide (1) was dissolved in 1 liter of dry CH 2 Cl 2, followed by the addition of 16.2 ml (0.2 mol) of pyridine and 29.8 g (0.2 mol) of trichloroethanol. Then 20 g (0.1 mol) of dicyclohexylcarbodiimide were added and the mixture was stirred for 16 hours at 220. Dicyclohexylurea began to precipitate; after precipitation was completed, the urea was removed by filtration. The filtrate was washed with 200 ml of a 5% aqueous solution of sodium bicarbonate, 200 ml of 10% phosphoric acid and 100 ml of a saturated aqueous solution of sodium sulfate. The organic phase was dried over sodium sulfate at 50 for 30 minutes, filtered and evaporated to an oil. Dietary ethers were added and, while scraping, product 2 crystallized out in an amount of 27 g (yield 63%).

Step 2. + 3 Quinoin IN 26.5 g (0.062 mol) of compound 2 were dissolved in 500 ml of p-dioxane and then 8.5 ml (0.078 mol) of benzoic chloride and 8.75 ml (0.078 mol) were added. ) kinoiin. The solution was refluxed for 4 hours and then dissolved in 300 ml of water and the product 3 was extracted with 2 x 400 ml of ethyl acetate.

The ethyl acetate extracts were combined, washed successively with 200 ml of a 5% aqueous solution of sodium bicarbonate, 200 ml of 5% phosphoric acid and 200 ml of a saturated aqueous solution of sodium sulfate, dried over sodium sulfate at 5 ° for 30 minutes and evaporated to dryness. an oil (3), which was used as such in the subsequent reaction. 49 466 203 Step 3 2 ", + mou + 112o2 1 glacial acetic acid '0 0 l / -íT / I // Cæg u, rN« «' <> o '_ É The compound obtained in the previous step was dissolved in 1 liter of glacial acetic acid. p and "under * omröriffngß vidta-ZZP * ti-Tlsattëšfšdroppvisff'ttadw vatnïöšrfingzav fi ze.gf_f __. l'r ~ potassium permanganate until a light red color remained (ie a drop placed on a piece of filter paper) gave a light red color. Then 30% hydrogen peroxide was added dropwise with cooling until a clear solution was kept; a certain amount of white precipitate was present. The solution was poured into 2.5 liters of water and the product 4 was extracted with 3 x 500 ml of ethyl acetate. The ethyl acetate was washed with a 5% aqueous solution of sodium bicarbonate to a neutral reaction (ie until some bubbles no longer appeared on addition), dried over sodium sulfate and evaporated to give the remaining product 4. This was allowed to stand for 1 day at 10 ° and then triturated with "Skellysolve B" to give 9.1 g of the solid product 4. The yield was 28% of the theoretical value for steps 2 and 3 in combination. 466 205 50 Step 4 L; "'1 glacial acetic acid + ®KEH o o \\ // nagon.

S m / C 5 (BL-Paola) Hz l, __ N å o 2 c (See US-P 4 164 497) 3; 7 $ '9' zinc dust slurried f '5 “mF” šätüikàfoc fi “É? Tdes“ tiTT ”5 To this mixture was added a solution of 3 g (0.0057 mol) of compound 4 in 15 ml of dimethylformamide and the resulting slurry was stirred for 2.5 hours at 5 °. The zinc was then filtered off and the pale yellow solution was poured into 80 ml of a 5% aqueous solution of hydrochloric acid. The mixture was extracted with 3 x 25 ml of ethyl acetate. The combined ethyl acetate extracts were extracted with 3 x 20 ml of a 5% aqueous solution of sodium bicarbonate, the ethyl acetate phase being recovered after separation.

The bicarbonate extracts were combined, introduced under a layer of ethyl acetate, adjusted to pH 1.5 by the addition of 2N hydrochloric acid and saturated with sodium sulfate. The ethyl acetate was separated and the aqueous phase was extracted with 2 x 30 ml of ethyl acetate. All of the above ethyl acetate phases were combined, dried over sodium sulfate and evaporated to an oil (consisting of the free acid form of BL-P 2 O 13), which was dissolved in about 20 ml of acetone, to which 20 ml of diethyl ether was then added. Then 50% potassium -2-ethylhexanoate (KEH) in dry n-butanol was added to neutral reaction. The product si 466 205 (BL-P20l3) crystallized out. After stirring for 0.5 hours at 220 DEG, the product was collected by filtration to give 650 mg of the product 5 (37% yield).

A 50 mg sample of product 5 was dissolved in 0.5 ml of water and 20 mg of N, N'-dibenzylethylenediamine (DBED) diacetate was added. The DBED salt of product 5 crystallized out, was collected by filtration, washed with water and dried over P 2 O 5 in vacuo to give N, N'-dibenzylethylenediamine-2beta-chloromethyl-2-methylpenam-3-carboxylate sulfone ( The DBED salt of the free acid 5).

Another sample of product 5 (450 mg) was dissolved in 3 ml of water and to the solution was added a solution of 270 mg of DBED diacetate in 2 ml of water. During scraping, the DBED salt of product 5 (430 mg) crystallized out. Recrystallization from about 5 ml of boiling acetone gave 270 mg.

Example 23 6 - [(R) -2-Amino-2-p-hydroxyphenylacetamide] -3,3-dimethyl-7-oxoph4-thia-1-aza-bicyclo [3.2] 2-heptane] -2-carbonyloxymethyl-2-beta-chloromethyl-2-alpha -methylphenamine .. »-3alpha-carboxylate sulfone of the formula wa IHD CIí - 430 - NH¶., - S, _ I Qkcíäâ _ I _ _N J” f NI-Iz o I c C = 0 vil »0 -o esf / ci-Lzcl C 1 ”C33 I .l- fl 466 203 æ was prepared by replacing the ampicillin used in the procedure of Example 21 with the corresponding Dane salt of amoxycillin.

BIOLOGICAL DATA The product 5 according to Example 1 of the formula 'qäí / ° cnzcl' f "w COZK will hereinafter be designated BL-P2013.

Although the compound BL-P2013 in itself is a very weak antibacterial agent, the compound inhibits beta-lactamases and protects ceforanide and amoxicillin from attack by beta-lactamase-producing bacteria in vitro and infvivoavíd fi use combination combination fi with β2 β2 activity. in BL-P2013 in comparison with ampicillin "IK 019m) Organism BL-P2013 Ampicillin. pneumoniae _ A-9585 16 0.004 B. pyogenes A-9604 63 0.004 lä: aureus A-9537> l25 0.16 S. aureus + 50% serum A -9537> l25 0.06 S. aureus Pen- Eaes A-ssoe> 12s> l25 S. aureus Meth- Res Al5097> l25 125 S. faecalis. A20688 => l25- ". 0.l3w§ n .. C611 »Alsns« S12? - i 'h' 'i' E. coli A2034ll25> l25 K. pneumoniae Al5l30> l25 125 K. pneumoniae A20468> l25> l25 P. mirabilis A-9900> l25 0.13 P. vulgaris A2l559> l25- _l2S P. morganii Al5l53> l25> l25 P. rettgeri A21203> l25 4 S. marcescens A200l9> l25 16 * E. cloacae A-9659> l25 63 E. cloacae A-9656> l25> l25 P. aeruginosa A ~ 9843A> l25> l25 P. aeruginosa A21213> l25> l25 466 205 ~ 54 Tabe11 2 Bacteroidal activity of ceføranide and amoxycíïïin separately and in combination with BL-P2013 _ 1261: 6- _ MIK (pslfmUš. _ ° F låšš ”ršzïšâ” íââfšâäïâ 333; 1192611111 êäa * in (l: l) (lzl) B. fragilis 1121916 + 63 2> 12s 2 a 1122053 * + 32 2 63 2 a 1122021 + '32 2 32 _2_ 4 1121375 + 32 2 63 2 a 1122534 +> 12s 22 125 ä> 12s 1122697 + 63 2 63 2 s 212269 + 63 _4_ 63 2 16 1122694 + 125 22 63 2 16 112269 +> 12s _1_6_ 32 2 125 E2269 f tvi;> l2S ; 1% '_§§g,; §fl ñ3 fi p' "3§¿¿_2§> I251.2, 22253 + 5”> 12s ”_32 * - 32" 32 "5 '112253 +> 12s 22 125 _3_2_> 12s 112279 +> 12s 2 32 _4_ 125 112279 + 32 2 32 2 s 112279 + 32 _4_ 32 2 s 212279 + 63 '2 32 _4_ 16 112279 + 63 2 63 2 16 112279 + 32 2 63 2 58 13. théta- iotaomíc- - _ H gon A222? + 125 2 63 2 16, 112227 + 125 2 63 2 16 55 466 203 Table 2 (continued) Anti-Bacteroides activity of cephoranide and amoxycillin separately and in combination with BL-P2013 -n- o-. em-_ MIK (fl s / mll * Organism läkt-f Cefo- Ceforanid MFP * Ämoxycill in Amoxvz amas rania + sL-p2 013 2013 + 3_z.-1 = 2o13 cinm '(l.:l) (1: l) Bacteroides -, stammar "" A2o93 + 32 _4¿ 32 2 s A219s + 63 ¿_ 32 2 16 22092 + ss _4_ 32 _2_ 16 A2l95 + 63 Lä 63 2 16 _ A209: + 63 _1_¿s_ 63 _ 5_ a A2093 + 125 å 125 g_ 32 "A209: + 63 5_ 32 _2_ 16 i ~ Azoszï- - 2o:; $ '= ;. 13.2. 63: (xml-if o' .." 13ï ~ f ~ ål A2093 4 2 2 125 0.132 0; I3 '- good synergism marginal synergism - * The minimum inhibitory concentration (-MIK) was measured by the agar dilution method using 50-fold dilution of 24-tin culture cultures as inoculum, which had been stored in a Steers inoculator. Brucella agar plus 5% leached sheep blood and 10 pg / ml vitamin K. 466 203 - 56 Table 3 Therapeutic activity of amoxycillin in combination with BL-P2013 when administered to mice infected experimentally with a beta-1-lactamase strain of Staphylococcus aureus Infection dose PD50 / treatment (mg / kg) Organism (an number of organisms) Amoxycillin (A) BL-P2013 (B) A + B (1: 1) IM P0 IM P0 IM P0 s. aureus sx 108> soo åsoo _> so> zoo 6.3 44 A-9606 5 x 108> 800> 800> 50> 200 19 77 7 x m8.> soo -.> so - 9.5 - Schedule of treatment: the compounds were administered 0 and 2 hours after infection.

Table 4 Blood levels of BL-P2013 and its pivaloyloxymethyl ester (BL-P2024) after oral administration.tñJH? NiE fi â¿g¿¿mi .. _ Blood level (pg / ml) u. Dose Half-Test Association (mg / kg) 15 30 60 90 120 150 ring time orga_ Minutes after administration (min.) Nism 100 4.5 5.1 3.9 2.3 1.5 0.8 50 E. coli A-9675 ßL-Pzois wo 4.6 4.1 3 <2.6 <2.6 <2.s - s. Aureus A-9606 200 7.4 9.8 7. ~ 1 4.1 2.8 42.6 50 _._- S. aureus * 'A-9606 100 12.8 12.9 9.7 7.2 5.5 4.2 70 E. coli A-9675 BL-P2024 100 13.1 12 8 5.7 3.8 (2.5 60 S. aureus A-9606 200 14.7 14.4 9.8 8.7 5.1 <2.5 60 S. aureus _ A-9606 The values given in the table are the mean values of 2-4 experiments 57 466 205 Tabe11 5 Blood levels and delivery times for BL-P2024 after ear1 administration of different doses to mice Biod level (pg / ml) Compound gas / k) is so so so izø iso r 1/2 mg 9 Minutes after administration (min.) 5.9 6.2 3.6 1.9 1.3 0.7 40 BL-P2024 50 7.7 9.5 6.3 4.7 3; 5 2.3 60 100 12.3 12.2 9.4 7.2 5.7 4.6 85 200 14.7 14.4 9.8 8.7 5.1 <š2.5 60 BL-P2024 was suspended in Tween-CMC water.

The values given in the table are the average values of 2 trials at the dosage levels 25 and 50 mg / kg, of 5-6 trials at the dosage level of 100 mg / kg and of 2-3 trials at the dosage level of 200 mg / kg.

Test organism: E. coli A9675 at all dose levels with the exception of 200 mg / kg of BL-P2024 (Sa auneus.A9606). 466 203 QONA m.n ~ ß QQNA mo fl X ß _ coN ^. , v.. nv fl nv m flfl om. ooN ^ noe X m mßncm fi vß fi wßum fl vvnnm mN “mN Nnuev vvue fi w. ooNA maa X w m fl wu fl m .m OONA NmA «ww oo fi noc fl ßmuv fifi Ü ooæA maa X m Hmom fl <°° ~ ^« «" «v mn fl wß @@“ ~ ww¶ ° = ~ ^ »QX x« møuusn .m = @ ~ ^ ° ~, = H v fl uv fi ß fl nnn °° @ ^ QQH x ~ w ° w @ <== ~ ^ Qa fi ßmw wßumn nnnnn -. «V ° m ^. °° mMl ° = ~ ^ Qø fl x ß mswusø .w n fl c fl vad Nu fl fl u fi fl u fl anv 1 - I x @. = + ~ _Wu Åßwsm- = m Am n fl onm fl hm ”= ICuAxøE <| axoe <nøoßo 23.5: Em fl cmmuø 1. _ moum = o @« ¥ m »= - in .. twåmsv m5 = ë2wn .. \ o¶nnm * Foo .u zuo mnwgzm .w> ~ Lmssßvm wu = mLmu = uo; Qm @ E ~ wx fl pløumn UQE v ~ Pw ¥ CmEwLOQx0 UUGL | 0Hxw%: w mmm E - ~ p mzwgwguwwc m ~ oN @ | 4m uws: o @ ~ n = _nso ¥ W m -Pwpøh =? __ ~ u> xos ~> ~ »wpw> ~» xø xmwu = mq ~; w »466 203 .uzu fi W wumLwu = wQm : m Lm> øcgwmcwcugmm .zcwpxwwcw xwuww Løsswv N: oo o vpmgo wwvmgwgpmwzwsum ~: L ~ m: @ = wLm »uøsuzuwwmc * -ucwzwm ° = ~ ^ @ ~, m wm wuw n« ~ .- °° «. W == ~ ^ m¿m. ~ Ssaß. Nw n. ~ .E ~; °° ~ ^ mo fi x ß m ~ wm <°° ~ ^ w ~. «~ W« nwn nww ~. @ °° ~ ^ mø fi x Q flfi co .m -. .V \ n fi øu «. ~. ~» ~ «.H H. ~. .H.« _§W __ @ u> x ° E <^ L ~ Ew. =.

.Tum. m_oN.T._m: _.-_ Zu> .x ° s <.... ». .èmLo ÉÉS Em fi cnmuo, N. woum = ow »¥ mw = ~ ^ @ ¥ \ @Ev @ = * _ ¶ = ~ =« @ \ ° mnw_J_.

* Foo .m zuo m = mL: ~ .m> ~ Løeampm muc ~ @ mu = uo »Qw ~ sm» xø ~ | ~ uwn ams »~ _mp: we« LwQxo own; nmøzwwcw mmmë - ~ u mcwamxuwpcwsum w +> mo ~ @ | 4m Ume copuølwpëox v: @ - «uäxoEø> ø uw» w> waxø xmPu: waøLwh Ü ~ I | III | IIII .møgow Q -mnøH é0 466 203 @? _ m @ @ _ L ~> =? __ Wu> x ° E <.omza & mm «? ~ @@ _ m @ _ L ~> ß = L ~ = ° w - =? @ E ° ¥ - <~ ° ~ @ -4m "= p__Pu> x ° E <@, @ ° @ p ¥ w» = WL @ ~ »@ Lßeawp N Sug 0» _ ~ L ° ~ u ~ _u = ~ = @@ wmwzß .wg fl gwn ~ == mn P cowmcmqmaw som_wu m; wL ~ wP = ~ sv ~ «~ oN @ | 4m: mums ^ ømo _ = - Pwu-æuwewxopgmx - = «@ = h @w» _ß == @ »-> V 0: 0» W.; m ~ w «<°° ~ ^>.> m. ~.>«. ~ “~ .M °° Q», uzua ma fl xw fifi oo .m _ L »w> w @ <mw ^ www mun ~" @ °° @ ~ uzoa mø fi x ß flfi øu .m _ ... omzo mßwmc _. m ~ A qnw m.m “> m. ~” ° H °° »~» cm mø fl x ß fifl ou .m. ; "omzn mßwmm mw A. man nn» ~ H ”~ H Qc fl k» om maa xm flfi co .m _ B »« ß @ ° ~ <°° ~ ann @, ~ H QQW. uzoa øø fl xm unwusß .m _ _ . ~ @ = m fi <°° ~ ^ fi mußm nnuww = ° @ w @ ozua @ ° fi. xm mswuø fl .m ”_ vwow fl. H Au fl fi uv = @ W ~ wv n = ww = m» m: = m Agmsmwc _ mLun vuoumlam »E. ~ Eu> xos <..>. xos <2 Pwškm: mv -mono Évcß Eu fl cømuø _ .i ._ x: mLøL:. ß n @CW _. Om. Gå w ° Üm = ° HP . ¥ @ *> »-% w ~ OU .w: OO w3UL: ß.m> ø LøE @ = wL @ L @ m * = PEw ~ vv, ~ _o ~ @ -4 = w = @ _ ~ @ o »Xosø> ~» m »~>« u ¥ ~ xmwøzunøgwh | s ~ ßm ww = ~ »mu = uo; Qmßs ~ ux- | ~ pw @ was» -w »= ms * Lunxw, mu ~ Lwp ¥ w + = P mwms -pwp> ~ ^ «NoN @ -4mv =; w» mw ~ »» ms @ xo-> o ~ ø> ~ Q vws Lmzopßufwßaox mxwpo ~ ~ _m; ß »466 203 ^ -P = ¥ ° = P »_. =" M. @ N = Qv _ V m ~ wm <@_ou .u Uswmcmmgouwwh »mp ->» uwmWw = wu «» = o ¥ «mw | wmm» mn »mm> ~ ww» = wL ~ Q soc * ~ = wu »m> -â = w» »m>» uzu | = wwz »W ~ u__w« ms ~ L »ø =; mm = ~ = w; wm á / ^ mw ~ MH ~ ^ n . = w + mHv Amwmmwv ^ w.m fl m% w.mww »nV ^ #» mwH.mv oo fl n ~ omm »nm: fl w fl wdv Qám fi â Swww flfi v G.M4H_A ~ aw fi wwwwv Cmwmwäv om Doäàm w.ov w.ov w.ov_ ^ m.%@w.ov iw.H «w» ¶v ^ ww «mH.Hv mw n ~ omm- @ m Smmwdv Sáw flfi. Quwmmi: fi w fl mxï Sßw fi wm, âwmwæv 02 wå fi z fi m w.ov ^ m. ~ Wmmo. ^ :. ~ m + »ov ^ nm ~ m.HV ~ ^ nwmm« v ^ww@mw.~v_ om wno «m-Am w.ov w.ov m.øv. ^ nw ~ m.ov> @ ^ wmwmmv ^ nww «n.Hv mm wnowm | qm. @ = @ LmL ~ m« = «su ~» L @ w + w gmïzcwz øm fl om fi .om Seša: Lå fi ïšwm fi m on ma wuzßms m fi cuàï F xnmßmm mmmwmwwmw » m.uw> vmoNm | 4m: oo m ~ oNm | 4m .mmo ~ @ | 4m> ø aww> *: uo ~ mm ~ _wnø ». ^ »~ _ = ¥ ° N_.o“ @. @ U: av m ~ mm <W-ou.m Usmwcmmgo fi wmh? M = wu * w = ox mmw | Nmm »mn Lww> ~ mw» = mß ~ Q soc * ~ = wuLw>: w ~ u ~> | u: u «m wa»: oo Poxzpmcwpzaogn am W av-muwemgw ucgømzwzwgmm 62 466 205 Amßš, 2%: s fi mwmmv a fi fn å åšåm QWWS Gamww .. G fi mm mv v: nâ w .ov ^>. Hm @ wov ^ w. ~ w ++ ov ^ ww ~ w.Hv .M mn mm wnowm-qm Awwmå âøwwwmv Cuwm fi nv Ankmqwav Sm fi mm fi v 03 .aomnïqm SWWS: Émmw fl v: H fifi mc: fi w fi m fl RAM fi. om âowïhm wo ^ m.ov ^æ.~m@wov ^ æ.m «m.ov Wwmwwnv ^ mM @ ww.« v mm = mo «mL» m @ = * »w &« m * = * E¶ ~ ; @p »@ gw» == ~ = om fl ¶ 93 ¶ om om. om 3 Awåww fi 9.222 ~ _E \ @ = v Lw «> * = @ Q_ = N xmwgmm 466 203 ^ -. = ¥ ° N_. ° u @. @ U: av m ~ mm <W-øu.u" sw @ = øm; o »mw» »wF ~ @> ßmwmvm =« u? »= o ¥ ~ m« | wmm »mv Lwm> ~ mwpcmgøn sea * ~ = wuLw> = w ~« ~> | u: u | = um: »zuo ~ o ¥ æ ~ m = w ~ aQoL = um w mwp-mumsmgw øcgømcpcwßmu fw .. 63 m xmmxmm ^ @ fl w. fl ~ ^ wm-w.« v Mwwnv ^m.wm@wmv ^ @@ mW ~ w.nV% M oo fi wnowmfqm ^ mm ~ w.ov ^ n | «.Hv ^ = mwmHv Abwww fl v @ w¶w ~ w.nv ^ Nw ~ w.mV om wnomm | qn Awnwl fi v ^ m. = | Hv ^ æ.: wmwHv ^ ~.> w + wHv fi w@m~m.mv ^ mm ~ w. «v mm wno ~ m | qm ^ m.m fl w.ov ^ w.æm« »~ v ^ æ.mHwwwnv % fi m »w»> v ^ w.m fl m. = v co fi nHowm »qm ïwm fl s fi mwmms cnmuumv: mwwmwv anwa om nñšaæ.

Andwwwov Sïmm fi ov Awwæmwnwï fi v ^ m. * Fi ~ ...% .. fi v mw nHommLHm @ = * Lw; »m * = @ su ~ Luuww Lwwfmrz A .m _ _w ¥ <@ w = v šv: om .466 203 54 Thus, the compounds of the present invention are useful for improving the efficacy of beta-lactam antibiotics with respect to beta-lactamase-producing bacteria upon oral and parenteral administration. On a weight basis, the dosage is from 1/5 to 5 times and preferably equal to the dosage of beta-lactam antibiotics. By way of example, the compounds of the present invention, in a ratio of 1: 1, markedly enhance the activity of cephoranide and amoxycillin with respect to beta-lactamase-producing strains of anaerobic Bacteroids, such as B. fragilis, B. thetaiotaomicron and other species belonging to that genus. and also with respect to resistant strains of Staphylococcus aureus. The compounds of the present invention are administered either in admixture with or concomitantly with the beta-lactam antibiotic in a dosage within the stated ratio of the known and conventional dosage of the antibiotic.

Thus, the ability of the compounds of the present invention to enhance the efficacy of a beta-lactam antibiotic with respect to certain beta-lactamase-producing bacteria makes them valuable for co-administration with certain beta-lactam antibiotic agents.Treatment of the bacterial infection ~ in ~ mammalian gurus: specieñtsgjuhosem of a bacterial infection, a compound of the present invention may be mixed with the beta-lactam antibiotic and the two agents may then be administered simultaneously. Alternatively, a compound of the present invention may be administered as a separate agent during treatment with a beta-lactam antibiotic.

When using a compound of the invention or a salt thereof. to enhance the antibacterial activity of a beta-lactam antibiotic, the compound may be administered as such or preferably in a formulation together with standard pharmaceutical carriers and diluents. A compound of the present invention, which is in the acid form or as a pharmaceutically acceptable salt thereof, may be administered orally or parenterally; a compound of the present invention in the form of an ester which is readily hydrolyzed in vivo is best administered orally. Parenteral routes of administration include intramuscular, subcutaneous, intraperitoneal and intravenous administration. When a compound of the present invention is used in the presence of a carrier or diluent, the carrier or diluent is selected with respect to the intended mode of administration. For example, if one intends to administer the compound orally, it may be used in the form of tablets. capsules, lozenges, powders, syrups, tinctures and mixtures, aqueous solutions and aqueous suspensions and the like, in accordance with pharmaceutically standardized practice. The ratio of active ingredient to carrier is, of course, dependent on the chemical nature, solubility, stability and potency of the active ingredient as well as on the intended dosage. However, these pharmaceutical compositions usually contain 5-80% of carriers. In the case of tablets for oral use, commonly used carriers include substances such as lactose, sodium citrate and salts of phosphoric acid. Various disintegrants, such as starch, and lubricants, such as magnesium stearate, sodium lauryl sulfate and talc, are commonly used in tablets. For oral administration in capsule form, useful diluents are lactose and high molecular weight polyethylene glycols. When aqueous suspensions are required for oral use, the active ingredients are combined with emulsifying and suspending agents. If desired, certain: sweeteners and / or flavors, may be added, parenteral, administered; which, including: intramuscularly, intraperitoneally, intramuscularly and intravenously, usually sterile solutions of the active ingredient are prepared and the pH of the solutions is adjusted and buffered appropriately. For intravenous use, the total concentration of soluble substances should be checked to make the preparation isotonic.

Although the physician treating the patient ultimately determines the dosage to be applied to the patient in question, the ratio between the daily dosage of a compound of the present invention or a salt thereof and of the beta-lactam antibiotic is usually from 1: 5 to 5: 1 and preferably 1: 1. In addition, the daily oral dosage of each component is normally l0-200 mg per kg body weight and the daily parenteral dosage of each component is normally l0-100 mg per kg body weight. These values are illustrative only and in some cases it may be necessary to use dosages outside the specified limits.

Claims (2)

466 205 46 PATENT CLAIMS Esters of the formula 32Cl 324% S --1 * 'C33 _ É'OR wherein R is benzyl or substituted benzyl. A process for the preparation of a compound according to claim 1, wherein there is at least eat x fi, at & nman_uQgQettar, a formerly eningïmed «forme1n ~ N --_. ° 4L- 4? C-OR || Wherein R is benzyl or substituted benzyl, in an inert, anhydrous organic solvent in the presence of large, equimolar amounts of a weak tertiary amine and an acid chloride, until the reaction is substantially complete.