Process for preparing new derivatives of cephalosporine

(42) Date of commencement of the patent: 15.10.1986 (45) Date of issue: 25. 5.1987 (30) Priority:

29th 3.1982 US 363313 claims.

(73) Patent owner:

BRISTOL-MYERS COMPANY NEW YORK (US).

(54) PROCESS FOR THE PREPARATION OF NEW CEPHALOSPORIN COMPOUNDS scar

TOR 0073018

Nr.383129

The invention relates to a process for the preparation of the novel compounds of the general

formula

<img file="AT383129B_D0001.tif" />

wherein R¹ represents a hydrogen atom or a conventional amino-protecting group, R² and R³ each 5 is independently a methyl or ethyl group and R is⁴ a methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, allyl, 2-butenyl, 3-butenyl, 2-hydroxyethyl,

3-hydroxypropyl, 2- (dimethylamino) ethyl, pyridylmethyl, pyridylethyl, benzyl or Phenäthylgruppe means and their non-toxic, pharmaceutically acceptable acid addition salts or solvates.

, GB Patent No. 1,399,086 comprises a large number of cephalosporin compounds of the general

formula

<img file="AT383129B_D0002.tif" />

wherein R represents a hydrogen atom or an organic group, R^a an etherified, monovalent, organic group bound to the oxygen atom via a carbon atom,

B is ^&gt; S or ^ 5S - »- 0 and P is an organic group. However, the 2-aminothiazol-4-yl group is not among the R substituents. There are also no examples in which P is described as the quaternary ammonium methyl group as in the compounds of the present invention. Similar disclosure has been found in US Pat. No. 3,971,778 and Expt. Nos. 4,024,133, No.4,024,137, No.4,064,346, No. 4,033,950, No. 4,079,178, No. 4,091,209.

No. 4,092,477 and No. 4,093,803 thereof.

U.S. Patent No. 4,278,793 broadly describes a large number of cephalosporin derivatives of the general formula

<img file="AT383129B_D0003.tif" />

wherein the variables R¹, R², R³, R⁴, X and A are the general definitions of the corresponding 25 substituents of the compounds of the formula (I) claimed in the invention. In the 20 columns in which the different substituents are defined, in the 78-page

However, it is not described that A can be a quaternary ammonium methyl group. British Patent No. 1,604,971 is the British equivalent thereto and essentially describes the same facts. GB Patent No. 2,028,305 A, apparently unrelated formally, has the same broad disclosure content, but A means only one hydrogen atom.

Nr.383129

U.S. Patent No. 4,278,671 describes 7- [2- (2-aminothiazol-4-yl) -2- (syn) -methoxyiminoacetamido] cephalosporin derivatives of the general formula

<img file="AT383129B_D0004.tif" />

wherein R²NH is an optionally protected amino group and R³ represents a hydrogen atom or the residue of a nucleophilic compound. The term residue of a nucleophilic compound is broadly defined to find that R³ Alternatively, it may be a quaternary ammonium group. Dabie only pyridinium, variously substituted pyridinium, quinolinium, picolinium and lutidinium are described as quaternary ammonium groups. However, there is no indication that the quaternary ammonium group of those in the inventive

Compounds can match. British Patent No. 1,581,854 is the British equivalent thereto and has substantially the same disclosure. Other patents of the same assignee, which are formally unrelated but of similar disclosure, are U.S. Patent Nos. 4,098,888 and US Patent Nos. 4,203,899, 4,205,180 and 4,298,606, and GB PS No. 1,536,281.

GB-PS No. 2,040,921 describes cephalosporin derivatives of the general formula

<img file="AT383129B_D0005.tif" />

<img file="AT383129B_D0006.tif" />

wherein R^a and R^b represent a C 1-4 alkyl group or, together with the carbon atom to which they are attached, form a C 3-7 cycloalkylidene ring and R¹, R² and R³ each represents a C j_4-alkyl group.

The preparation of compounds of the general formula (I) is explained in the following reaction scheme.

The abbreviation Tr means the trityl (triphenylmethyl) group, which is a preferred amino-protecting group. The abbreviation Ph stands for the phenyl group. The - [CH (Ph)₂ ] Radical is thus the benzhydryl group, which is a preferred carboxyl-protecting group.

<img file="AT383129B_D0007.tif" />

(II)

- 4 No.383129 (III)

<img file="AT383129B_D0008.tif" />

(IV) (V)

<img file="AT383129B_D0009.tif" />

(VI) (VII)

<img file="AT383129B_D0010.tif" />

(I) the compound (VII)

Process produced

The crucial reaction step in the reaction scheme is the reaction with the tertiary amine. The compound (VII) can also be obtained by others.

Nr.383129

The inventive method for the preparation of compounds of general formula (I) and their non-toxic, pharmaceutically acceptable salts and solvates is accordingly characterized in that a compound of the formula

<img file="AT383129B_D0011.tif" />

wherein R² has the meanings given above, B^l a common carboxyl protecting group and B²a conventional amino protecting group, with a tertiary amine of the general formula

<img file="AT383129B_D0012.tif" />

wherein R³ and R⁴ have the meanings given above, to a compound of the general formula

<img file="AT383129B_D0013.tif" />

(XV) and then removing all protecting groups in a manner known per se and / or, if desired, converting the compounds of formula (I) into a non-toxic, pharmaceutically acceptable salt or solvate.

Preferred compounds of the general formula (I) which are: 7 - [(Z) -2- (2-aminothiazol-4-yl) 15 -2-methoxyiminoacetamido] -3- (trimethylammonium) methyl-3-cephem-4-carboxylate ( la), 7- [(Z) -2- (2-

-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- [N, N -dimethyl-N- (2-hydroxyethyl) ammonium] methyl-3-cephem-4-carboxylate (lb), 7- [(Z) -2- (2-aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- (N, N-dimethyl-N-allylammonium) methyl-3-cephem-4-carboxylate (Ic), 7- [(Z) -2- (2-aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- [N, N -dimethyl-N- (3-pyridylmethyl) ammonium] methyl-3-cephem-4-carboxylate (Id),

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- [N, N-dimethyl-N- (2-dimethylaminoethyl) ammonium] methyl-3-cephem-4 carboxylate (Ie), 7- [(Z) -2- (2-aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- (N, N-dimethyl-N-ethylammonium) methyl-3-cephem-4 carboxylate (If) or 7- [(Z) -2- (2-aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- (N, N-diethyl-N-methylammonium) methyl-3-cephem -4- carboxylate (Ig) and its non-toxic, pharmaceutically acceptable acid addition salts or solvates are prepared according to the invention so that benzhydryl 3-iodomethyl-7- [(Z) -2-methoxyimino-2- (2-tritrylaminothiazol-4-yl) acetamido] -3-cephem-4-carboxylate with Trimethylamine, N, N-dimethyl-N-2-hydroxyethylamine, N, N-dimethyl-N-allylamine, 3- (dimethylaminomethyl) pyridine, 1,2-bis (dimethylamino) ethane, dimethylethylamine or diethylmethylamine to give the corresponding quaternary salt deblocking quaternary salt and / or if desired, the resulting compounds are converted to a non-toxic pharmaceutically acceptable acid addition salt or solvate.

Another embodiment of this process for the preparation of 7 - [(Z) -2- (2-aminothiazol-4-yl) -2-ethoxyiminoacetamido] -3- (trimethylammonium) methyl-3-cephem-4-carboxylate is that Benzhydryl 3-iodomethyl-7- [(Z) -2-ethoxyimino-2- (2-tritylaminothiazol-4-yl) acetamido] -3-ce6

No. 3883129 phem-4-carboxylate with trimethylamine, deblocked the quaternary salt and / or, if desired, the resulting compound in a non-toxic, pharmaceutically acceptable acid addition salt or solvate transferred.

The reaction is carried out in a nonaqueous organic solvent such as methylene chloride, chloroform, ethyl ether, hexane, ethyl acetate, tetrahydrofuran, acetonitrile and the like or in mixtures of such solvents. The reaction is conveniently carried out at a temperature of about -10 to about + 50 ° C; Preferably, the reaction is carried out at room temperature. At least 1 mole of tertiary amine should be used per mole of compound (XIV); Preferably, about 50 to 100% excess tertiary amine is used.

For the above reaction as B¹ useful carboxyl protecting group are known in the art and include aralkyl groups such as benzyl p-methoxybenzyl, p-nitrobenzyl and diphenylmethyl (benzhydryl); Alkyl groups such as tert-butyl; Haloalkyl groups, such as 2,2,2-trichloroethyl, as well as further, in the literature, for example in GB-PS No. 1,399,086, described carboxyl protecting groups. It is preferable to use carboxyl-protecting groups which can be easily removed by treatment with acid, particularly preferred carboxyl-protecting groups are the benzylhydryl and tert-butyl radicals.

As B² useful amino protecting groups are also known and include, for example, the trityl group and acyl groups such as the chloroacetyl group. Amino-protective groups which can be easily removed by treatment with acid, for example the trityl group, are preferred.

The tertiary amines of the formula used in the process according to the invention are used

<img file="AT383129B_D0014.tif" />

are commercially available or can be readily prepared in a manner known per se.

The invention also includes the preparation of the solvates (including the hydrates) of the compounds of the general formula (I) and of the tautomeric forms of the compounds of the formula (I), for example the 2-iminothiazolin-4-yl form of the 2 -Aminothiazol-4-yl radical.

As can be seen from the structural formula, the compounds of formula (I) may be in syn or Z configuration with respect to the alkoxyimino group. Since the compounds represent geometric isomers, they may also partially exist in the form of the anti-isomer. The invention includes compounds of formula (I) which contain at least 90% of the syn isomer. Preferably, the compounds of formula (I) are syn-isomers which are substantially free of the corresponding anti-isomers.

The non-toxic, pharmaceutically acceptable acid addition salts of the compounds of general formula (I) include the salts, for example with hydrochloric acid, hydrobromic acid, formic acid, nitric acid, sulfuric acid, methanesulfonic acid, phosphoric acid, acetic acid and trifluoroacetic acid, and with other acids which on the penicillin and Cephalosporin area were used.

The compounds of the formula (I) wherein R is¹ is a hydrogen atom, have a great antibacterial activity against various Gram-positive and Gram-negative bacteria and are useful for the treatment of bacterial infections in humans and animals. The compounds of formula (I) may be formulated for parenteral use in a conventional manner using known pharmaceutical carriers and excipients. They may be presented in unit dosage form at a level of preferably from about 50 to about 1500 mg of active ingredient of formula (I) in multiple dose containers. If desired, the compounds obtainable according to the invention can be administered in combination with further antibiotics, such as penicillins or other cephalosporins.

The dosage in adults is usually in the range of about 500 to about 5000 mg per day, depending on the frequency and mode of administration. For intramuscular or intravenous administration to adults, a total dosage of approximately

Nr.383129

- 7 750 to about 3000 mg per day in divided doses normally sufficient, although higher daily doses of some of the compounds obtainable according to the invention may be desirable in the case of Pseudomonas infections.

The preferred compounds of formula (I) are those wherein R¹ a hydrogen atom 5, R² a methyl group, R³ a methyl or ethyl group and R⁴a methyl, ethyl, 2-hydroxyethyl, 2- (dimethylamino) ethyl, allyl or pyridylmethyl group. Particularly preferred compounds are those wherein R¹ a hydrogen atom, R² and R³ a methyl group and R⁴ a methyl, 2-hydroxyethyl or allyl group. The most preferred compounds are R¹ for a hydrogen atom and R², R³ and R⁴ represent a methyl group. For evaluation of the preferred compounds, the minimum inhibitory concentrations (MIC) of the compounds were determined by the two-series agar dilution method in Mueller-Hinton agar against 32 strains of test organisms in six groups. The geometric mean MIC values ​​obtained in this test are summarized in Table 1.

Table 1

<td rowspan="2">Connection of the example</td><td colspan="6">Geometric mean of MIC values ​​(μg / ml)</td>

<td>(G +) - Ia (5 strains)</td><td>(G +) - Ib (5)</td><td>(G -) - Ia (5)</td><td>(G -) - Ib (6)</td><td>(G -) - II (5)</td><td>(G -) - III (6)</td>

<td>_x(A)</td><td>1.1; 1.6</td><td>3.6; 3.6</td><td>0.033; 0,029</td><td>0.25; 0.20</td><td>0.69; 0.69</td><td>3.2; 4.0</td>

<td>2</td><td>1.6</td><td>4.7</td><td>0,038</td><td>0.18</td><td>0.60</td><td>3.5</td>

<td>3</td><td>1.4</td><td>3.6</td><td>0,044</td><td>0.22</td><td>0.69</td><td>5.6</td>

<td>4</td><td>2.1</td><td>4.7</td><td>0.057</td><td>0. 22</td><td>0.69</td><td>7.1</td>

<td>8th</td><td>2.1</td><td>5.5</td><td>0.087</td><td>0.50</td><td>1.4</td><td>7.9</td>

(G +) - Ia: penicillin-sensitive S. aureus (5 strains) (G +) - Ib: penicillin-resistant S. aureus (5 strains) (G -) - Ia: cephalothin-sensitive E. coli (2 strains), Pneumoniae (1 strain) and Pr. Mirabilis (2 strains) (G -) - Ib: cephalothin resistant E. coli (3 strains) and Kl. Pneumoniae (3 strains) (G -) - II: Pr. Morganii (1 strain), Ent. Cloacae (2 strains) and Ser. marcescens (2 strains) (G -) - III: Ps. aeruginosa (6 strains)

Two different batches of compound (la)

The absorption of the most preferred compound (Ia) (prepared according to Example 1) was determined in mice following a single intramuscular injection of the test compound (dissolved in 0.1 M phosphate buffer; p ^ 7) at a dose of 20 mg / kg. Blood samples were taken from the orbital sinus, placed in heparinized capillary tubes and tested in Mueller-Hinton20 medium using Morganella Morganii A9695 as a test organism. The blood level values ​​at different times, the value for the half-life (t 1) and the area under the curve (AUC) are summarized in Table 2.

- 8 No.383129

Table 2

Blood levels of compound (Ia) in mice

<td>blood levels</td><td>10</td><td>14</td>

<td>(Pg / ml)</td><td>20</td><td>12</td>

<td>min</td><td>30</td><td>8.8</td>

<td>to</td><td>40</td><td>7.5</td>

<td>administration</td><td>50</td><td>4.7</td>

<td></td><td>60</td><td>4.4</td>

<td></td><td>90</td><td>1.5</td>

<td></td><td>120</td><td>0.74</td>

<td>⁴1.2</td><td></td><td></td>

<td>(Min)</td><td></td><td>24</td>

<td colspan="2">AUC gg.h / ml</td><td>10</td>

The in vitro activity of the most preferred compound (Ia) against 31 strains of difficult-to-control bacteria was determined in GC agar. The results are summarized in Table 3.

Table 3

In vitro activity of compound (Ia) against difficult-to-control bacteria

<td>test organism</td><td>Geometric mean of the MIC values (Gg / ml)</td>

<td>S. pyogenes (6 strains)</td><td>0,013</td>

<td>S.pneumoniae (6)</td><td>0,013</td>

<td>N. gonorrhoeae (4)</td><td>0,013</td>

<td>N. meningitidis (5)</td><td>0.016</td>

<td>H. influenzae (7) (Ampicillin sensitive)</td><td>0,013</td>

<td>H. influenzae (3) (Ampicillin resistant)</td><td>0.20</td>

The following examples illustrate the invention, with the prepended preparation examples relating to the preparation of starting compounds according to the above reaction scheme.

Production Example 1

N ·

-COOCjHg (III)

TrHN '

OR²

Äthy 1- (Z) -2-methoxyimino-2- (2-tritylaminothiazol-4-yl) acetate (purple)

A mixture of ethyl (Z) -2-hydroxyimino-2- (2-tritylaminothiazol-4-yl) acetate (II) (5.0 g;

10.0 mmol) CHjJ (2.04 ml, 32.8 mmol) and K₂CO₃ (4.54 g, 32.8 mmol) in dry dimethylnr.383129

- 9 sulfoxide [(DMSO) 100 ml] is stirred overnight at room temperature and then in

Water (250 ml) poured. The precipitate formed is filtered off, washed with water and dried to give the title compound (5.15 g, quantitative yield). Melting point 115 ° C (decomposition).

NMR: «CDC1₃ ppm: 1.32 (3H, t), 3.98 (3H, s), 4.30 (2H, q), 6.42 (1H, s), 7.2 (1H, m),

7.25 (15H, s).

Compound (Illb) is prepared according to the above general procedure but using ethyl iodine instead of methyl iodide.

<td>connection</td><td>R²</td><td>Yield (%)</td><td>Mp (° C)</td><td>literature ^ Mp (° C)</td>

<td>(purple)</td><td>methyl</td><td>100</td><td>115 (Zers.)</td><td>about 120 (Zers.)</td>

<td>(IIIb)</td><td>ethyl</td><td>67</td><td>97-98</td><td>*)</td>

*) The ester is hydrolyzed without isolation Tetrahedron, 34, 2233 [1978]

Production Example 2

C-COOH (IV)

TrHN ^ OR² (Z) -2-Methoxyimino-2- (2-tritylaminothiazol-4-yl) acetic acid (IVa)

The ethyl ester (purple) (6.0 g, 12.7 mmol) prepared in Preparation Example 1 in ethanol (120 ml) is treated with 2N NaOH (12.7 ml) overnight at room temperature. The

Reaction mixture is added by adding powdered dry ice to p_ß 8 set. The solvent is evaporated under reduced pressure. The residue is dissolved in water (100 ml) and the solution is acidified to pH with 1N HCl_ß 2 and then extracted with ethyl acetate (3 χ 50 ml). The combined extracts are washed with a saturated aqueous NaCl solution, dried and evaporated. The residue is crystallized from ethyl acetate-hexane to give 5.56 g (yield 987o) of the title compound [mp 138-143 ° C (dec.)]. NMR: δ CDC1₃ ppm: 3.89 (3H, s), 6.52 (1H, s), 7.2 (15H, s).

Compound (IVb) is prepared according to the above general procedure.

<td>connection</td><td>R²</td><td>Yield (%)</td><td>Mp (° C)</td><td>literature Mp (° C)</td>

<td>(IVa)</td><td>methyl</td><td>98</td><td>138-143</td><td>about 140</td>

<td>(IVb)</td><td>ethyl</td><td>85</td><td>140-145</td><td>not specified</td>

⁽¹⁾ Tetrahedron, 34, 2233 [1978]

Production Example 3

Benzhydryl-7-amino-3-chloromethyl-3-cephem-4-carboxylate (V)

To a slurry of PC1₅ (8.3 g, 40 mmol) in CH₂C1₂ (100 ml) is added pyridine (3.2 g, 40 mmol) and the mixture is stirred for 20 min at 20 ° C. Benzhydryl-3-hydroxymethyl-7-phenylacetamido-3-cephem-4-carboxylate (5.1 g, 10 mmol) is added to this mixture with stirring at -40 ° C. at once. The mixture is stirred for 15 min at -10 ° C and allowed to stand for 7 h at -10 to -15 ° C. To the cooled solution (-20 ° C) is added propane-l, 3-diol (10 ml) and the mixture is 16 h at -20 ° C and then stirred for 20 min at room temperature. The resulting solution

Wash with ice-water (2 times 20 ml) and saturated aqueous NaCl (10 ml) over MgSO 4₄dried and concentrated in vacuo. The gummy residue (12 g) is dissolved in a mixture of CHCl₃ and n-hexane (2: 1) and chromatographed using a silica gel column (200 g) and using the same solvent as the eluent. Those fractions containing the title compound are evaporated in vacuo and the residue is triturated with n-hexane to give the title compound (2.1 g, 51%) with a melting point of 110 ° C (dec.).

IR: v KBr 3400, 2800, 1785, 1725 cm¹,

UV: λ ^Et0H 265 nm (E ^ 160).

max 1 cm

NMR: δ DMSO-d₆ + CDC1₃ ppm 3.69 (2H, s), 4.43 (2H, s), 5.09 (1H, d, J = 4.5Hz), 5.24 (1H, d, J = 4.5Hz) , 6.87 (1H, s), 7.3 (10H, m).

Production Example 4

Benzhydryl-3-chloromethyl-7- [(Z) -2-methoxyimino-2- (2-tritylaminothiazol-4-yl) acetamido] -3-cephem-4-carboxylate (Via)

Benzhydryl-7-amino-3-chloromethyl-3-cephem-4-carboxylate prepared according to Preparation 3 (2.29 g, 5.52 mmol) in CH₃CN (57 ml) is treated with bis (trimethylsilyl) acetamide (BSA, 4.09 ml, 16.6 mmol) at room temperature for 50 min to give a clear solution. To this solution is added an acid chloride solution prepared from (Z) -2-methoxyimino-2- (2-tritylaminothiazol-4-yl) acetic acid (IVa) (2.04 g, 4.60 mmol) and PClg (1.15 g, 5.52 mmol) in methylene chloride (20 ml). The mixture is stirred at room temperature for 30 minutes, poured into cold water (200 ml) and extracted with ethyl acetate (3 × 100 ml). The combined extracts are washed with aqueous NaCl, dried and evaporated. The syrupy residue (4 g) is chromatographed on a silica gel column (150 g) by eluting successively with 10: 1 and 3: 1 mixtures of toluene and ethyl acetate. Those fractions containing the desired compound are combined and the solvent evaporated to give 2.61 g (68%) of the compound (Via) in the form of an amorphous powder.

NMR: δ CDC1₃ ppm 3.50 (2H, s), 4.02 (3H, s), 4.34 (2H, s), 4.98 (1H, d), 5.87 (1H, q), 6.65 (1H, s), 6.90 (1H, s), 7.3 (25H, m).

Production Example 5

Benzhydryl-3-iodomethyl-7- [(Z) -2-methoxyimino-2- (2-tritylaminothiazol-4-yl) acetamido] -3-cephem-4-carboxylate (VIIa)

A mixture of the 3-chloromethyl derivative prepared according to Preparation 4 (Via) (1.50 g, 1.79 mmol) and NaI (1.34 g, 8.93 mmol) in methyl ethyl ketone (30 mL) is stirred at room temperature for 1 h , After evaporation of the solvent, the residue is dissolved in ethyl acetate (100 ml) and washed with water, aqueous Na₂S₂O₃ and aqueous NaCl, dried and evaporated to give the title compound (VIIa) (1.47 g, 89%) as an amorphous powder.

NMR: OCDC1₃ ppm 3.55 (2H, ABq), 4.00 (3H, s), 4.25 (2H, s), 4.97 (1H, d), 5.80 (1H, q), 6.65 ( 1H, s), 6.90 (1H, s), 7.3 (25H, m).

Production Example 6

Benzhydryl-3-chloromethyl-7- [(Z) -2-ethoxyimino-2- (2-tritylaminothiazol-4-yl) acetamido] -3-cephem-4-carboxylate (VIb)

To a solution of (Z) -2-ethoxyimino-2- (2-tritylaminothiazol-4-yl) acetylic acid (IVb) (1.095 g, 2.4 mmol) in dichloromethane (20 ml) is added phosphorus pentachloride (500 mg). After stirring at room temperature for one hour, the mixture is added all at once to an ice-cooled solution of compound (V) (1.083 g, 2.4 mmol) and BSA (1 mL) in dichloromethane (20 mL). After 0.5 hours of stirring, the reaction mixture is poured into 10% aqueous NaHCO 3₃ (200 ml) and extracted with CHCl₃ (100 ml). The extract is washed with water, dried over MgSO 4₄, and evaporated under reduced pressure. The residue is chromatographed on a silica gel column. Elution with CHC1₃ gives (VIb) as an amorphous powder; 1.76 g (86%).

NMR: δCDClj ppm 1.40 (3H, t, CH₂CH₃), 3.53 (2H, ABq, 2-CH₂), 4.37 (2H, s, -CH₂C1), 4.60 (2H, q, -CH₂CH₃), 4.90 (1H, d, 6-H), 5.89 (1H, d, 7-H), 6.88 (1H, s, thiazole-H), 6.91 (1H, s, benzhydryl -CH).

Nr.383129

Production Example 7

Diphenylmethyl-7- [(Z) -2-ethoxyimino-2- (2-tritylaminothiazol-4-yl) acetamido] -3-iodomethyl-3-cephem-4-carboxylate (VIIb)

A mixture of (VIb) prepared according to Preparation 6 (1.07 g, 1.25 mmol) and NaI (562 mg, 2.75 mmol) in acetone (20 mL) is stirred for 1 h. The mixture is filtered and the filtrate is poured into water and extracted into ethyl acetate. The organic layer is washed successively with 5% aqueous Na₂S₂O₃, Water and saturated aqueous NaCl, dried over MgSO 4₄ and evaporated to obtain 1.04 g (89%) of the compound (VIlb).

NMR: SCDClj ppm 3.55 (2H, q, 2-CH₂ ), 4.27 (2H, s, CH_ZD, 5.02 (1H, d, 6-H), 5.87 10 (1H, d, 7-H), 6.68 (1H, s, thiazole ring H), 6.93 (1H, s, benzhydryl -CH).

Example 1:

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- (trimethylammonium) methyl-3-cephem-4-carboxylate (Ia)

An IM solution of trimethylamine in diethyl ether (1 ml, 1 mmol) is added with stirring to a suspension of benzhydryl-3-iodomethyl-7 - [(Z) -2-methoxyimino-2- (2-tritylaminothiazole-4-) yl) acetamido] -3-cephem-4-carboxylate (VIIa) (468 mg, 0.5 mmol) in diethyl ether (30 ml) and the mixture is stirred for 1.5 h. The precipitated quaternary salt (XVa) is filtered off (410 mg, 82% yield), trifluoroacetic acid (TFA) (3 ml) is added. This mixture is stirred for 1.5 h at room temperature and then evaporated to dryness under reduced pressure below

20 ° C. The residue is triturated with ether and the precipitated TFA salt is filtered off (yield 365 mg), dissolved in a small amount of methanol and chromatographed on a column of HP-20 resin (1.8 x 20 cm). The column is eluted with water (about 1 liter) and then with 30% aqueous methanol (0.5 liter). The methanolic eluate is evaporated under reduced pressure below 40 ° C. The residue is lyophilized to give the crude title compound 25 (yield 129 mg). The ratio Δ ²/ δ³Isomers in the crude product is 1: 2, determined by HPLC. The product is purified by HPLC [Lichrosorb RP-18, (Waters Associates, Milford, Mass. USA.) 8χ300 mm, eluted with 1 / 100M NH₄H₂-PO₄ (p_H 7.2): CHjOH = 85:15].

The HPLC eluate is chromatographed on a column of HP = 20 (Mitsubishi, Tokyo, Japan) (1.8 x 15 cm) to remove inorganic salts. The column is eluted with water (0.5 l) and then with 30% aqueous methanol (0.5 l). The methanolic eluate is evaporated under reduced pressure below 40 ° C. The residue is freeze-dried to give the title compound (Ia) as an amorphous powder. Yield 75 mg [33%, based on (VIIa)]. The product gradually decomposes above 160 ° C. Purity about 80%.

IR: υ^KBr 3600-3000, 1775, 1660, 1610, 1540, 1350, 1030 cm "¹, τη 3.x

UV: λ phosphate buffer _{7) nm (ε) 2}35 (15700), 257 (15400).

NMR: «D₂O ppm 3.25 [9H, s, N⁺(CH₃)₃], 4,10 (3H, s, OCH₃), 5.47 (1H, d, 4Hz, 6-H),

5.96 (1H, d, 4Hz, 7-H), 7.10 (1H, s, thiazole-H).

Example 2

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- [N, N-dimethyl-N- (2-hydroxyethyl) ammonium] methyl-3-cephem-4 carboxylate (lb)

The general procedure described in Example 1 is repeated except that instead of the trimethylamine used therein, the equimolar amount of N, N-dimethylethanolamine is used. The crude product has a Δ² : A³Ratio of 1: 2. After purification, the title compound, which decomposes above 160 ° C, in 17% yield. Purity about 90%.

Example 3

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- (N, N-dimethyl-N-allylammonium) methyl 1 -3-cephem-4-carboxylate (lc)

The general procedure described in Example 1 is repeated except that an equimolar amount of N, N-dimethylallylamine 50 is used instead of the trimethylamine used therein. The crude product has a Δ: Δ ratio of 1: 4.5. After purification, the title compound, which decomposes above 150 ° C, in 14% yield. Purity about 80%.

Nr.383129

Example 4

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- [N, N-dimethyl-N- (3-pyridylmethyl) ammonium] methyl-3-cephem-4 carboxylate (Id)

The procedure described in Example 1 is repeated except that an equimolar amount of 3- (dimethylaminomethyl) pyridine is used in place of tri-methylamine. The crude product has a Δ²: Δ³Ratio of 1: 4.3. After purification, the title compound is obtained, which decomposes above 170 ° C in a yield of 17%. Purity about 75%.

Example 5:

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- [N, N-dimethyl-N- (2-dimethylamino-10-ethyl) -ammonium] -methyl-3-cephem-4 carboxylate (Ie)

The general procedure described in Example 1 is repeated except that an equimolar amount of 1,2-bis (dimethylamino) ethane is used instead of trimethylamine. The crude product has a Δ²: Δ³Ratio of 1: 1. After purification, the title compound, which decomposes above 150 ° C, in a yield of 14%. Purity about 65%.

Example 6

Z- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- (N, N -dimethyl-N-ethylammonium) methyl-3-cephem-4-carboxylate (If)

The general procedure described in Example 1 is repeated except that instead of the trimethylamine, an equimolar amount of Ν, Ν-dimethylethylamine is used.

The crude product has a Δ²: Δ³Ratio of 1: 1. After purification, the title compound, which decomposes above 150 ° C, in a yield of 15%. Purity about 77%.

Example 7

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- (N, N-diethyl-N-methylammonium) methyl-3-cephem-4-carboxylate (lg)

The general procedure described in Example 1 is repeated except that an equimolar amount of diethylmethylamine is used instead of trimethylamine. The crude product has a Δ²: Δ³ Ratio of 1: 1. After purification, the title compound, which decomposes above 150 ° C, in a yield of 10%. Purity about 65%.

Example 8

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-ethoxyiminoacetamido] -3- (trimethyl-ammonium) -methyl-3-cephem-4-carboxylate (Ih)

The procedure described in Example 1 is repeated except that instead of benzhydryl, 3-iodomethyl-7- [(Z) -2-methoxyimino-2- (2-tritylaminothiazol-4-yl) acetamido] -3-cephem-4 is used carboxylate (VIIa) used an equimolar amount of the corresponding ethoxyimino compound (VIIb) (prepared according to Preparation Example 6). The crude product has a Δ²: Δ³Ratio of 3: 1. After purification, the title compound, which decomposes above 150 ° C, in a yield of 3%. Purity about 70%.

Spectral Data for the Compounds of Examples 2 to 8 (a) Infrared Spectra (KBr)

All products give similar infrared spectra: 1770-1775 cm^-1 (β-lactam C = O), 1660 cm^-1 (CONH), 1610 cm^-1 (COO^-).

(b) ultraviolet spectra (1/15 M phosphate buffer; 7)

All products except those of Example 4 had similar spectra: 235 nM (e 15700-16400), 257 nM (ε 15400-16000). The compound of Example 4 had the following values:

235 nM (ε 17600), 255 nM (ε 18600, sh), 260 nM (ε 19000), 266 nM (ε 17900, sh).

(c) NMR spectra (D₂O)

- 13 - No.383129

<td rowspan="2">connection the Examples No.</td><td colspan="6">Chemical shift (ppm)</td>

<td>n⁺ch₃(S)</td><td>o-ch₃(3H, s)</td><td>6-H (1H, d, 4-5 Hz)</td><td>7-H (1H, d, 4-5 Hz)</td><td>Thiazole-H (1H, s)</td><td>other</td>

<td>2</td><td>3.20 (3H) 3.25 (3H)</td><td>4.10</td><td>5.46</td><td>5.90</td><td>7.10</td><td></td>

<td>3</td><td>3.08 (3H) 3.15 (3H)</td><td>4.10</td><td>5.46</td><td></td><td>7.10</td><td>5.6-6.4 (4H, m, 7-H, ch = ch₂)</td>

<td>4</td><td>3.04 (3H) 3.21 (3H)</td><td>4.09</td><td>5.45</td><td>5.94</td><td>7.06</td><td>4.75 (2Η, s, Pyr-CH₂) 7.66 (1H, dd, 8 &amp; 4 Hz, Pyr-H) 8,12 (1H, d, 8 Hz, Pyr-H) 8.74 (2H, s, Pyr-H)</td>

<td>5</td><td>3.18 (3H) 3.25 (3H)</td><td>4.10</td><td>5.45</td><td>5.90</td><td>7.10</td><td>2.50 [6 H, s, N (CH₃)₂]</td>

<td>6</td><td>3,10 (3H) 3,15 (3H)</td><td>4.10</td><td>5.45</td><td>5.90</td><td>7.10</td><td>1.48 (3H, t 7 Hz, CH₂CH₃)</td>

<td>7</td><td>3.05 (3H)</td><td>4.10</td><td>5.44</td><td>5.95</td><td>7.10</td><td>1.45 (6H, t, 7 Hz, CH₂CH₃)</td>

<td>8th</td><td>3.25 (9H)</td><td>-</td><td>5.50</td><td>5.97</td><td>7.10</td><td>1.45 (3H, t, 7 Hz, CH₂CH₃)</td>

It was found that the Δ²: A³And the product yield can be improved by allowing the filtrate of the originally precipitated quaternary salt (XV) to stand for a short time and again filtered through a bed of the originally obtained compound (XV);

in particular, the yield is improved by repeating the re-filtration several times. The yield can also be increased slightly by adding a small amount of the original tertiary amine to the filtrate. This will be explained with reference to the following example, which is a variation of Example 1.

Example 9

7- [(Z) -2- (2-Aminothiazol-4-yl) -2-methoxyiminoacetamido] -3- (trimethylammonium) methyl-3-cephem-4-carboxylate (Ia)

A solution of trimethylamine (1M solution in ether, 10 ml) is added all at once to a solution of iodine compound (VIIa) (4.68 g, 5 mmol) in ether (500 ml) with stirring. The mixture is stirred for 10 minutes and the precipitated quaternary salt (XVa) is filtered off and washed with a small amount of ether. The filtrate and the washings are combined and allowed to stand for a further 10 minutes at room temperature. The second precipitate is sucked off through the same filter on which the first precipitate is stored in the form of a bed. The filtrate is filtered three times in the same manner every 10 minutes to give the quaternary salt (XVa) [3.58 g; 72% from (Vlla)]. To the last filtrate is added a trimethylamine solution (1 M solution in ether, 2 ml) and the same filtration is repeated a further three times at 10 min intervals to give further (XVa) (0.56 g, 11%). To the obtained filtrate is added again an IM trimethylamine solution (1 ml) and the filtration is repeated twice at 10 minute intervals to obtain a third charge of (XVa) (0.134 g, 2.7%).

Nr.383129

The overall yield of (XVa) is 4.27 g (86%).

A mixture of (XVa) (4.20 g, 4.22 mmol), anisole (1 mL) and TFA (40 mL) is stirred for 1.5 h at room temperature. The mixture is evaporated under reduced pressure below

20 ° C and triturate the dark residue with isopropyl ether (300 mL) to precipitate the TFA salt (3.50 g), which is filtered off and dried under reduced pressure. Dissolve the TFA salt in methanol (50 mL), treat with a small amount of charcoal and filter. Concentrate the filtrate under reduced pressure and add to the concentrates sodium 2-ethylhexanoate [1M solution in ethyl acetate, 15 ml. Dilute the mixture with ethyl acetate (300 ml)] to give the crude title compound (2.36 g, purity about 50%). It is filtered off, washed with a small amount of ethyl acetate and dried. The ratio of Δ²Isomer to A³Isomer of the crude product is 1: 4 (HPLC, Lichrosorb RP-18, mobile phase, 1 / 100M ammonium phosphate buffer, ρ ^ 7 - CHjOH, 90:10; retention time, A²Isomer 6 '54, i³Isomer 8 '29).

The crude product (2.36 g) is dissolved in a small amount of water and purified by HPLC (Waters Associates, Milford, Mass. USA, System 500, PrepPAK 500 / C_l8 ; mobile phase, 7%

Methanol). The product containing HPLC eluate is concentrated under reduced pressure below 35 ° C. The concentrate is freeze-dried to give the title compound in an yield of 959 mg [42%, based on the iodide (VIIa)] as an amorphous powder. Estimated purity about 80% (by HPLC). The ratio of A²Isomer to A³Isomer is 1:17. The compound gradually decomposes above 160 ° C.