GB2049688A

GB2049688A - Cephalosporin Derivatives, Processes for the Preparation Thereof, and Antibacterial Compositions Containing Them

Info

Publication number: GB2049688A
Application number: GB8011651A
Authority: GB
Original assignee: Eisai Co Ltd
Current assignee: Eisai Co Ltd
Priority date: 1979-04-09
Filing date: 1980-04-09
Publication date: 1980-12-31
Also published as: FR2453865A1; CH645905A5; NL8002083A; FR2453865B1; GB2049688B; DE3013675A1; IT8021260A0

Abstract

New cephalosporin derivatives useful as antibacterial drugs and represented by the general formula: <IMAGE> wherein R<1>, R<2>, R<3> each represents hydroxy, acyloxy, and hydrogen provided that two or three of R<1>, R<2> and R<3> are not hydrogen; and R<4> represents hydrogen or hydroxy; and non-toxic salts thereof can be prepared by acylation of the corresponding phenylgycine or p- hydroxyphenylglycine cephalosporin using the appropriate optionally activated chromone carboxylic acid. They can be incorporated into pharmaceutical compositions.

Description

SPECIFICATION Cephalosporin Derivatives, Processes for the Preparation Thereof, and Antibacterial Compositions Containing Them This invention relates to novel cephalosporin derivatives.

The compound represented by the general formula (all):

wherein R4=H, is known as cephaloglycin and is disclosed in Belgian Patent No. 635,137 (1964). The compound represented by the general formula (II), wherein R4=OH, is also known, and is disclosed in British Patent No.1,240,687(1971).

The object of this invention is to provide novel antibacterial drugs.

The present inventions provides cephalosporin derivatives represented by the general formula (I):

wherein R1, R2, and R3 each represents a hydroxy, acyloxy, or hydrogen substituent, provided that not more than one of R1, R2, and R3 is a hydrogen, substituent, and R4 represents hydrogen or hydroxy; and their non-toxic salts.

Illustrative examples of acyloxy groups suitable for R', R2 and R3 in the compounds (I) of this invention include alkanoyloxy such as acetoxy or propionyloxy, and aroyloxy such as benzoyloxy.

Illustrative examples salts of compounds having the general formula (I) include those with inorganic cations such as sodium, potassium, calcium or ammonium, and those with organic cations such as triethylamine, dicyclohexylamine or procaine.

The compounds of this invention can be prepared by acylating the amino group in the phenylglycine orp-hydroxyphenylglycine moiety of the two known compounds (II).

Thus, the compounds of the general formula (I) of this invention can be prepared by a process provided by the invention which comprises reacting a compound of the general formula (Il), or a salt or hydrate thereof, with a compound represented by the general formula (Ill):

wherein R1, R2, and R3 are as set forth above, or with a reactive derivative of a compound (Ill).

When the carboxylic acid of the general formula (Ill) is itself used in the process, it is preferable to effect the reaction in the presence of a condensation reagent such as N,N'-dicyclohexylcarbodiimide, N,N'-diethylcarbodiimide, N,N'-diprnpylcarbodiimide, N-cyclohexyl-N'-morpholinoethylcarbodiimide, trialkyl phosphite, ethyl polyphosphate, phosphorus oxychloride, oxalyl chloride, or the like condensation agent.Examples of suitable reactive derivatives of the carboxylic acids of the general formula (Ill) include, for instance, an acid halide such as an acid chloride or bromide; a symmetric anhydride; a mixed anhydride such as one derived from a chlorocarbonate ester, trimethylacetic acid, thioacetic acid, diphenylacetic acid or the like other compound; a reactive ester such as one derived from 2-mercaptopyridine, cyanomethanol, p-nitrophenol, 2,4-dinitrophenol, pentachlorophenol, or the like other compound; a reactive acid amide such as N-acylsaccharin, N-acylphthalimide, as well as other derivatives.

The reaction between a compound of the general formula (11) and a compound of the general (Ill) can be conveniently carried out in an inert solvent at a temperature range of -50 to 500C, preferably at a temperature between -20 and 30 C, in the presence or absence of a basic or silylating reagent.

Illustrative inert solvents include acetone, tetrahydrofuran, N,N-dimethylacetamide, dioxane, dichloromethane, chloroform, benzene, toluene or ethyl acetate, and mixtures thereof.

Illustrative basic reagents include, for example, alkali hydroxides such as potassium hydroxide or sodium hydroxide; alkali hydrogen carbonates such as potassium hydrogen carbonate or sodium hydrogen carbonate; and amines such as triethylamine, pyridine or N-methylmoropholine. Illustrative silylating reagents include, for example, N,O-bis(trimethylsilyl)acetamide or hexamethyldisilazane.

The chromone carboxylic acid (Ill) used as the starting material in this invention can generally be obtained by oxidizing an aldehyde of the general formula (IV);

wherein R1, R2, and R3 are aciloxy or hydrogen substituents, as set forth above, with a Jones reagent (see Reagents for Organic Synthesis, Vol. 1, p. 142).

The chromone carboxylic acids wherein any of R1, R2 and R3 are hydroxy groups are obtained by such an oxidation, oxidizing an aldehyde (IV) which contains acyloxy group(s) in place of the desired hydroxy group(s), followed by hydrolysis of the unwanted acyloxy group(s). Selective deacetylation or some other procedure may be required where the acid (Ill) is a mixed acyloxy/hydroxy substituted compound.

The chromone carboxylic acid halides of the compound (Ill) can be obtained by reacting the acid of the formula (Ill) with a haIogenting reagent such as phosphorus pentachloride, thionyl chloride, etc.

The aldehyde of the formula (IV) can be prepared by the conventional processes, for example, the processes described in Tetrahedron 30, 3553 (1974).

Alternatively, the compound of the general formula (I), wherein R1, R2, and R3 each is hydroxy or hydrogen provided that two or three of R1, R2, and R3 are not hydrogen at the same time, of this invention can be prepared by hydrolyzing the acyloxy moieties of the compound (I) wherein R1, R2, and R3 each is acyloxy or hydrogen provided that two or three of R1, R2, and R3 are not hydrogen at the same time.

The alkali hydrogen carbonates such as sodium hydrogen carbonate, potassium hydrogen carbonate, etc. may be used for the purpose of effecting the said hydrolysis.

Illustrative of specified compounds of this invention are the following compounds and their sodium salts.

7,B-[D-2-(6,7-Diacetoxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3- cephem-4-carboxylic acid.

7j5[D-2-(6,7-Diacetoxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3- acetoxymethyl-3-cephem-4-carboxylic acid.

7ss-[D-2-(6,7-Dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3cephem-4-carboxylic acid.

7ss-[D-2-(6,7-Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic acid.

7ss-[D-2-(7,8-Dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3 cephem-4-carboxylic acid.

7ss-[D-2-(7,8-Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3 acetoxymethyl-3-cephem-4-carboxylic acid.

7ss-[D-2-(7,8-Dihydroxychromone-3-carboxamido)-2-phenylacdetamido]-3-acetoxymethyl-3 cephem-4-carboxylic acid.

7p-[D-2-(7,8-Dihydroxychromone-3-ca rboxamido)2-(4-hydrnxyphenyl)acetamidoj-3- acetoxymethyl-3-cephem-4-carboxylic acid.

7ss-[D-2-(6,8-Dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3 cephem-4-carboxylic acid.

7ss-[D-2-(6,8-Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3 acetoxymethyl-3-cephem-4-carboxylic acid.

7p-[D-2-(6,8-Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)aceta mido]-3- acetoxymethyl-3-cephem-4-carboxylic acid.

7ss-[D-2-(6,7,8-Dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3cephem-4-carboxylic acid.

7ss-[D-2-(6,7,8-Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3 acetoxymethyl-3-cephem-4-carboxylic acid.

7p-[D-2-(6,7,8-Trihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyí)acetamido]-3- acetoxymethyl-3-cephem-4-carboxylic acid.

The cephalosporin derivatives of this invention have excellent antibacterial activities and are effective not only against the Gram-positive bacteria, but also against the Gram-negative bacteria. The said cephalosporin derivatives are characterized in that they exhibit remarkable effect particularly, against Pseudomonas aeruginosa which cause hardly curable infections.

Moreover, it has been confirmed that the compounds of this invention show low toxicity in the toxicity test. For example, the acute toxicity values [LD50 (mouse, oral)j for the following compounds were over 5 g/kg: Sodium salt of 7,B-[D-2-(6,7-diacetoxychromone-3-carboxamido)-2-phenylacetamido]-3- acetoxymethyl-3-cephem-4-carboxylic acid; Sodium salt of 7,B-[D-2-(6,7-dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3- acetoxymethyl-3-cephem-4-carboxylic acid; Sodium salt of 7jB-[D-2-(6,7-diacetoxychromone-3-carboxamido)-2-(4- hydroxyphenyl)aceta mido]-3-acetoxymethyl-3-cephem-4-carboxylic acid;; Sodium salt of 7p-[D-2-(6,7-dihydroxychromone-3-carboxamido)-2-(4- hydroxyphenyl)acetamido]-3-acetoxymethyl-3-cephem-4-carboxylic acid.

Dosage of the compound of this invention as antibacterial drugs may range generally from about 2 to 300 mg/kg/day, and preferably from 10 to 100 mg/kg/day. The compounds of this invention can be administered orally or parenterally.

Pharmaceutical preparations containing the compounds of this invention can be produced by any conventional preparation processes. Therefore, this invention includes preparation compositions containing at least one of the present compounds suitable as the medicine for human. Such compositions are provided by conventional methods with any required pharmaceutical carriers or diluents.

Tablets and capsules for oral administration may be in unit dose presentation form, and may contain conventional excipients such as finding agents, syrup, gum arabic, gelatine, sorbitol, tragacanth or polyvinylpyrrolidone; diluents, for example, lactose, cornstarch, calcium phosphate, sorbitol, or glycine; lubricants, for example, magnesium stearate, talc, polyethylene glycol, or silica; disintegrants, for example, potato starch, or acceptable wetting agents, for example, sodium lauryl sulfate.

The said tablet may be subjected to coating by a process well known in the art.

Liquid preparations for oral administration may be in the form of aqueous or oily emulsion, solution, syrup, elixir, etc.

Alternatively, they may be dry products which can be redissolved in water or any suitable vehicle before administration.

Such liquid preparations may contain conventional additive(s), for example, a suspending agent such as sorbitol syrup, methylcellulose, glucose/sugar syrup, gelatine, hydroxyethylcellulose, carboxymethylcellulose, aluminum stearate gel and hydrogenated edible fat; an emulsifier such as lecithin, mono-oleic acid sorbitan and gum arabic, a non-aqueous vehicle such as almond oil, fractionated coconut oil, oily ester, propylene glycol and ethanol, an antiseptics such as methyl phydroxybenzoate, propyl p-hydroxybenzoate and sorbic acid.

The composition for injection is provided in an ampoule of unit dosage or in a vial with added antiseptics. The said composition may be in the form of suspension, solution, or emulsion in an oily or aqueous vehicle, and may contain formulating agents such as suspending agent, stabilizer and/or dispersant. On the other hand, the active ingredient may be powder which can be redissolved before administration in suitable vehicle, for example, sterile pyrogen-free water.

The preparations for injection can be provided using isotonic agents such as glucose, sodium chloride, sorbitol, etc., and if required, suspending agents, surfactants, pH controlling agents, etc.

Alternatively, the preparation for injection may be in the form of powder which can be dissolved before administration.

The suppository can be provided using a basis such as cacao butter, polyethylene glycol, Witepsol (trade mark, Dynamite-Nobel-Ag), etc., and, if required, a surfactant.

The following experiments and examples illustrate this invention, but are not to be construed as limiting the scope thereof.

Experiment 1 Preparation of Chromone-3-carboxylic Acids which are the Intermediates of the Compounds of this Invention A) 6,7-Diacetoxychromone-3-carboxylic Acid 6,7-Diacetoxychromone-3-carboxyaldehyde (1 7.8 g) was dissolved in 1 litre of acetone. To this solution was added with stirring Jones reagent (32.8 ml) which had been previously prepared by dissolving chromic acid (1 13.6 g) in concentrated sulfuric acid (1 15 ml) diluted with water to a volume of 500 ml.

The reaction mixture was concentrated to 100 ml, and poured into water (900 ml). The precipitates (6.5 g) were collected by filtration and recrystallized from ethyl acetate to obtain the desired compound (5.9 g).

B) 6,7-Dihydroxychromone-3-carboxylic Acid To 6,7-diacetoxychromone-3-carboxylic acid (1 5.3 g) produced in A) were added acetic acid (300 ml) and concentrated hydrochloric acid (100 ml), and the mixture was stirred for 20 minutes at about 700 C, and then cooled.

The precipitates were collected by filtration and recrystallized from dimethylformamide-water to obtain the desired compound (8.9 g).

7,8-Diacetoxychrnmoe-3-carboxylic acid and 7,8-dihydroxychromone-3-carboxylic acid were prepared by the processes according to A) and B), respectively. The properties of the resulting compounds are shown in Table 1.

Table 1

Elemental Analysis (%) Moleculer Ceiculated: (III) IR Spectrum Formula Found: No. R R R (cm-1, nujol) Melting point( C) C H 1 HO- HO- H 3370, 3300, 1730, 1635, 1620, C10H6O5 54,08 2.72 > 300 54.06 2.60 2 CH3COO- CH3COO- H 1780, 1760, 1730, 1620, C14H10O8 54.91 3.29 186-188 54.95 3.08 3 H HO- HO- 3380, 3275, 1725, 1620, C10H8O6 54.06 2.72 265-270* 53.65 2.53 4 H CH3COO- CH3COO- 1780, 1780, 1740, 1625, C14H10O8 54.91 3.29 178-179 54.90 3.25 *decompsoltion Example 1 7ss-[D-2(6,7-Diacetoxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3 cephem-4-carboxylic Acid and its Sodium Salt a) 6,7-Diacetoxychromone-3-carbonyl Chloride A mixture of 6,7-diacetoxychromone-3-carboxylic acid (918 mg, 3 mmol), benzene (20 ml), thionyl chloride (260 ,ul, 3.6 mmol) and N,N-dimethylformamide (0.2 ml) was refluxed for 2 hours and the solvent was removed. Benzene (5 ml) was added and then evaporated. This was repeated twice to remove traces of thionyl chloride. The residual solid was collected to afford the desired compound.

IR spectrum (cm-l, nujol): 1780,1775,1660,1625.

b) 7ss-[D-2-(6,7-Diazetoxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3cephem-4-carboxylic Acid N,O-Bis-(trimethylsilyl)acetamide(1.483 ml, 6 mmol) was added to a stirred suspension of cephaloglycin (811 mg, 2.0 mmol) in dichloromethane (16 ml) at OOC. The mixture was stirred for 20 minutes at OOC, to which was added dropwise a suspension of the acid chloride (2 mmol) described in a) in dichloromethane (10 ml). The stirring was continued for 0.5 hour at OOC and for a further 5 hours at room temperature and the solvent was removed. The residue was taken up in ethyl acetate (250 ml), washed successively with 0.5 N hydrochloric acid (40 mlx2), water (40 mlx2) and saturated brine (40 mlx2) and dried (MgS04).After removal of the solvent, the residue was triturated with ethyl ether to afford the desired compound (920 mg, 66%).

Melting point: 170-1 750C (decomposition).

Elemental analysis: for C32H27N30,3S C H N Calculated (%): 55.41 3.92 6.06 Found(%): 54.38 3.86 5.05 IR spectrum (cm~1, nujol): 1780,1735,1690,1665, 1620.

c) Sodium Salt of the Compound Described in b) Sodium 2-ethylhexanoate (0.5 M solution in ethyl acetate, 2 ml) was added to a solution of the compound (694 mg, 1 mmol) described in b) in a mixture of acetone (30 ml) and N,Ndimethylformamide (2 ml). Ethyl acetate-ethyl ether (1:1, 30 ml) followed by ethyl ether (50 ml) was added to the mixture and the precipitate formed was filtered off, washed with ethyl acetate-ethyl ether (1:1) and dried to afford the desired compound (306 mmg, 43%).

Melting point: 165-1 800C (decomposition) Elemental analysis: for C32H26N3NaO13S C H N Calculated (%): 53.77 3.66 5.87 Found (%): 52.35 3.83 5.37 IR spectrum (cm~', nujol): 1 785, 1 770, 1 742, 1670, 1 605.

NMR spectrum (8, DMSO-d6): 2.00 (3 H, s), 2.37 (3 H, s), 2.38 (3 H, s), 3.13(1 H, d, J=1 7 Hz), 3.43 (1 H, d, J=1 7 Hz), 4.78 (1 H, d, J=12 Hz), 4.94 (1 H, d, J=4.5 Hz), 5.02 (1 H, d, J=12 Hz), 5.61 (1 H, dd, J=8 Hz, 4.5 Hz), 5.94(1 H, d, J=8 Hz), 7.25-7.7 (5 H, m), 7.92 (1 H, s), 8.1 2 (1 H, s), 9.09 (1 H, s),3.49 (1 H, d, J=8 Hz), 10.15(1 H, d,J=8 Hz).

Example 2 7ss-[D-2-(6,7-Diacetoxychoromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic Acid and its Sodium Salt a) 6,7-Diacetoxychromone-3-carbonyl Chloride A mixture of 6,7-diacetoxychromone-3-carboxylic acid (18.4 g, 60 mmol), benzene (450 ml), thionyl chloride (8.6 g, 72 mmol) and N,N-dimethylformamide (3 ml) was refluxed for one hour and cooled to room temperature. After the addition of n-hexane (300 ml), the resulting precipitate was filtered off to give the desired compound (17.6 g).

IR spectrum (cm~', nujol): 1780,1755,1660,1625.

b) 7ss-[D-2(6,7-Diacetoxychromone-3-carboxamido)-2-(4-hydroyphenyl)acetamido)-3acetoxymethyl-3-cephem-4-carboxylic Acid N,O-Bis(trimethylsily)acetamide (1 M solution in dichloromethane, 5 ml) was added to a stirred suspension of 7fi-[D-2-amino-(6,7)diacetoxychromone-3-carboxamido)-2-(4- hydroxyphenyl)acetamido]-3-acetoxymethyl-3-cephem-4-carboxylic acid (475 mg) in ethyl acetate (5 ml) at OOC. The mixture was stirred for 20 minutes at OOC, to which was added a solution of the acid chloride (325 mg) described in a) in dichloromethane (5 ml). After stirring for 4 hours at OOC, the reaction mixture was diluted with ethyl acetate, washed successively with water, 0.5 N hydrochloric acid, water and saturated brine, and dried (MgSO4).The solvent was evaporated and the residue was triturated with ethyl ether-n-hexane (2:1) to afford the desired compound (526 mg).

Melting point: 21 0-2200C (decomposition) Elemental analysis: for C32H27N30,4S C H N Calculated (%): 54.16 3.84 5.92 Found (%): 52.54 3.83 5.69 IR spectrum (cm-', nujol): 1760-1790,1735,1665, 1620.

NMR spectrum (#, DMSO-d6): 2.04(3H,s), 2.36(3H,s), 2,37(3H,s), 3.41(1H,d, J=18Hz), 3.57(1H,d,J=18Hz),4.67(1 H,d,J=13 Hz), 4,98(1H,d,J=13 Hz), 5.05 (1H, d, J=5Hz), 5.6-5.9(2H, m),6.75(2H,d,J=9Hz), 7.28 (2 H, d, J=9 Hz), 7.88 (1 H, s), 8.07 (1 H, s), 9.05 (1 H, s), 9.40(1 H, d, J=8 Hz), 10.00 (1 H, d, J=8 Hz).

c) Sodium Salt of the Compound Described in b) According to the method described in Example 4-c), the desired compound (379 mg) was obtained from the compound (497 mg) described in b).

Melting point: about 2300C (decomposition) Elemental analysis: for C32H26N3NaO14S C H N Calculated (%): 52.53 3.58 5.74 Found (%): 49.85 4.11 5.54 IR spectrum (cm-', nujol): 1720-1780,1665,1610.

Example 3 7ss-[D-2-(6,7-Dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3cephem-4-carboxylic Acid and its Sodium Salt a) 6,7-Dihydroxychromone-3-carbonyl Chloride A mixture of 6,7-dihydroxychromone-3-carboxylic acid (9.0 g, 4 mmol) and thionyl chloride (1 50 ml) was refluxed for 45 minutes and the thionyl chloride was evaporated. After the addition of benzene, the mixture was evaporated again to dryness and the residue was triturated with n-hexane to afford the desired compound (9.1 g, 93.4%).

IR spectrum (cm-1. nujol): 1780,1765,1645,1625.

b) 7ss-[D-2-(6,7-Dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-4carboxylic Acid N,O-Bis(trimethylsilyl)acetamide (44.8 g, 54.4 mml, 220 mmol) was added to a stirred suspension of cephaloglycin (16.2 g, 40 mmol) in dichloromethane (400 ml) at 20C. The mixture was stirred for 2 hours at 20C, to which was added the acid chloride (8.7 g, 36 mmol) described in a) and the stirring was continued for 1.5 hours at 20C. The reaction mixture was diluted with ethyl acetate (1 litre), washed successively with ice-cooled, saturated brine, an ice-cooled mixture of 0.5 N hydrochloric acid and saturated brine (4 times) and dried (MgSO4). The solvent was evaporated and the residue was triturated with n-hexane to afford a crude product (8.25 g).A suspension of a portion (1.8 g) of the product in acetone (60 ml) was stirred at room temperature for 2 days and after removal of acetone, the residue was triturated with ethyl ether and washed with n-hexane to afford the desired compound (1.5 g; Yield allowing for the untreated crude product: 28.2%).

Melting point: 240-2550C (decomposition) Elemental analysis: for C28H23N30"S C H N Calculated (%): 55.17 3.80 6.89 Found (%): 50.37 3.68 5.41 IR spectrum (cm-', nujol): 1780, 1735, 1710, 1660, 1630.

NMR spectrum (, DMSO-d6): 2.04 (3 H, s), 3.40 (1 H, H, d, J=18 Hz), 3.52(1 H, H, d, J=18 Hz), 4.70(1 H, H, d, J=12 Hz), 4.98(1 H, H, d, J=12 Hz), 5.03 (1 H, d, J=5 Hz), 5.74(1 H, H, dd, J=8 Hz, 5 Hz), 5.87 (1 H, d, J=8 Hz), 7.02 (1 H, s), 7.20-7.52 (5 H, m), 7.44 (1 H, s), 8.88 (1 H, S), 9.50 (1 H, H, d, J=8 Hz), 10.42 (1 H, d, J=8 Hz).

c) Sodium Salt of the Compound Described in b) Sodium 2-ethylhexanoate (0.5 M solution in ethyl acetate, 6.0 ml) was added dropwise to a solution of the compound (1.83 g, 3 mmol) described in b) in a mixture of acetone (850 ml) and N,Ndimethylformamide (13 ml). The mixture was stirred for 20 minutes at room temperature, to which was added successively ethyl acetate-ethyl ether (1:1, 800 ml) and ether (200 ml). After stirring for 30 minutes at room temperature, the precipitate was filtered off, washed with ethyl acetate and dried to afford the desired compound (1.24 g 65.4%).

Melting point: 185-1950C (decomposition) Elemental analysis: for C28H22N3NaO'1S C H N Calculated (%): 53.25 3.67 6.65 Found (%): 48.73 3.74 5.53 IR spectrum (cm~', nujol): 1765, 2740, 1660, 1630, 1610.

NMR spectrum (#, DMSO-d6): 2.01(3 H, 5), 3.20(1 H, d, J=18 Hz), 3.42 (1 H,d,J=18Hz), 4.80(1 H, d, J=12 Hz), 4.94(1 H, d, J=5 Hz), 5.01 (1 H, H, d, J=1 2 Hz), 5.59 (1 H, br.s), 5.90 (1 H, d, J=8 Hz), 6.80 (1 H, s), 7.20-7.60 (5 H, m), 7.29 (1 H, s), 8.76 (1 H, s), 9.43 (1 H, br.s), 10.64(1 H, H, d, J=8 Hz).

Example 4 7ss-[D-2-(6,7Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3 acetoxymethyl-3-cephem-4-carboxylic Acid and its Sodium Salt a) 6,7-Dihydroxychromone-3-carbonyl Chloride A mixture of 6,7-dihydroxychromone-3-carboxylic acid (888 mg, 4 mmol) and thionyl chloride (25 ml) was refluxed for one hour and the thionyl chloride was evaporated. After the addition of benzene, the mixture was evaporated again to dryness and the residue was triturated with dichloromethane to give the desired compound (719 mg).

IR spectrum (cm-', nujol): 1780,1765,1645,1625.

b) 7ss-[D-2-(6,7-Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic Acid N,O-Bis(trimethylsilyl)acetamide (742 yI) was added to a stirred suspension of 7ss-[D-2-amino-2- (4-hydroxyphenyl)acetamido]-3-acetoxymethyl-3-cephem-4-carboxylic acid (238 mg) in ethyl acetate (10 ml) at OOC. The mixture was stirred for 20 minutes at OOC, to which was added the acid chloride (120 mg) described in a). After stirring for 4 hours at room temperature, the reaction mixture was diluted with ethyl acetate, washed successively with water and saturated brine, and dried (MgSO4) and the solvent was evaporated.

Acetone was added to the residue and the mixture was allowed to stand at room temperature overnight. After removal of acetone, the residue was triturated with ethyl ether to afford the desired compound (101 mg).

Melting point: about 2500C (decomposition) Elemental analysis: for C32H23N30,2S C H N Calculated (%): 57.06 3.44 6.24 Found (%): 51.84 3.96 5.76 IR spectrum (cm-1, nujol): 1785, 1730, 1715, 1660, 1630, 1610.

NMR spectrum (#. DMSO-d6): 2.04 (3 H,s), 3.40 (1H, d, J=18 Hz), 3.58 (1 H, d, J=18Hz), 4.67 (1 H, d, J=13Hz), 4.99(1H, d, J=13 Hz), 5.05 (1 H, d, J=5 Hz), 5.64-5.88 (2 H, m), 6.74(2 H, d, J=8.5 Hz), 7.00(1 H, 5), 7.27 (2 H, d, J=8.5 Hz), 7.44 (1 H, s), 8.88 (1 H, s), 9.36(1 H, d, J=8 Hz), 10.28 (1 H, d, J=8 Hz).

c) Sodium Salt of the Compound Described in b) Sodium 2-ethylhexanoate (0.5 M solution in ethyl acetate, 0.24 ml) was added to a solution of the compound (80 mg) in acetone-N,N-dimethylformamide (2:1,3 ml). To the mixture was added ethyl acetate-ethyl ether (1:1, 10 ml).

The precipitate formed was filtered off, washed with ethyl acetate-ethyl ether (1 :1) and dried to afford the desired compound (67 mg).

Melting point: about 2350C (decomposition) Elemental analysis: for C32H22N3NaO,2S C H N Calculated (%): 55.25 3.19 6.04 Found (%): 50.03 4.06 7.04 IR spectrum (cm~', nujol): 1730-1780,1660, 1600-1630.

NMR spectrum (S, DMSO-d6-D2O): 2.04 (3H,s), 3.16(1H,d,J=18 Hz), 3.48 (1H,d,J=18Hz), 4.74(1H,d,J=13Hz). 4.90(1H.d.

J=13 Hz), 4.91(1 H, H, d, J=5 Hz), 5.60(1 H, H, d, J=5 Hz), 5.64(1 H, H, s), 6.80 (2 H, d, J=8.5 Hz), 7.03 (1 H, s), 7.31(2 H, H. d, J=8.5 Hz), 7.44(1 H, 5), 8.83 (1 H, s).

Example 5 7ss-[D-2-(6,7-Dihydroxychromone-3-carboxamido)-2-phenyliacetamido]-3-acetoxymethyl-3cephem-4-carboxylic Acid Sodium hydrogen carbonate (1 M solution in water, 0.698 ,ul) was added to a solution of the compound (200 mg, 0.279 mmol) described in Example 1-c) in water (160 ml). The mixture was allowed to stand at room temperature for 7 days, acidified to pH 2.0 and then extracted with ethyl acetate (50 ml x4). The organic layer was washed successively with water and saturated brine, dried (MgSO4) and evaporated.Trituration of the residue with ethyl ether afforded the desired compound (51 mg, 30%). This compound was identical with the compound described in Example 3-b) in all respects [NMR, IR and thin layer chromatography (silica gel, Merck 5715, benzene/dioxane/acetic acid=4:1 :1)].

Example 6 7ss-[D-2-(6,7-Dihydroxychromons-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic Acid Sodium hydrogen carbonate (1 N solution in water, 546 jul) was added to a solution of the compound (200 mg) described in Example 2-b) in water (15.45 ml). The mixture was allowed to stand at room temperature overnight, adjusted to pH 2.0 with 1 N hydrochloric acid and extracted with ethyl acetate. The organic layer was washed successively with water and saturated brine, dried (MgSO4) and evaporated. The residue was purified by thin layer chromatography on silica gel to afford the desired compound (108 mg).This compound was identical with the compound described in Example 4-b) in all respects [NMR, IR, and thin layer chromatography (silica gel, Merck 5715, benzene/dioxane/acetic acid=4:1 :1)].

Example 7 7ss-[D-2-(7,8-Diacetoxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3cephem-4-carboxylic Acid a) 7,8-Diacetoxychromone-3-carbonyl Chloride A mixture of 7,8-diacetoxy-3-carboxylic acid (9.5 g, 31 mmol), thionyl chloride (2.6 ml), N,Ndimethylformamide (0.1 ml) and benzene (300 ml) was refluxed for 1.5 hours. After the addition of thionyl chloride (2.6 ml) and N,N-dimethylformamide (0.1 ml), the mixture was refluxed for an additional hour and evaporated to dryness. The residue was triturated with n-hexane to afford the desired compound (9.3 g, 92.6%).

IR spectrum (cm-1, nujol): 1780,1770,1670,1620.

b) 7ss-[D-2-(7,8-Dlacetoxychromone-3-carbgoxamido)-2-phanylacetamido]-3-acetoxymethyl-3cephem-4-carboxylic Acid N,O-Bis(trimethylsilyl)acetamide (1 M solution in dichloromethane, 5.0 ml) was added dropwise to a stirred suspension of cephaloglycin (405 mg, 10 mmol) in ethyl acetate (10 ml) at OOC. The mixture was stirred for 1 hour at OOC, to which was added the acid chloride (325 mg, 1.0 mmol) described in a), after stirring for 4 hours at OOC, the reaction mixture was diluted with ethyl acetate (300 ml), washed successively with water (40 ml), 0.5 N hydrochloric acid (40 mlx2), water (40 mlx2) and saturated brine (40 mlx2) and dried (MgSO4) Removal of the solvent afforded pale yellow crystals (402 mg).A portion (108 mg) of the crystals was purified by preparative thin layer chromatography on silica gel (developing solvent: chloroform/methanol/formic acid=90:10:4) to give the desired compound (24 mg, yield allowing for the untreated crude product: 14.0%).

Melting point: 150-160 C (decomposition) Elemental analysis: for C32H27N3013S C H N Calculated (%): 55.40 3.92 6.06 Found (%): 54.64 4.10 4.85 IR spectrum (cm-', nujol): 1780, 1740, 1720, 1685, 1670, 1615, NMR spectrum (#, DMSO-d):: 2.03 (3 H, s), 2.38 (3 H, s), 2.44 (3 H, s), 3.42 (1 H, d, J=1 8 Hz), 3.52 (1 H, d, J=1 8 Hz), 4.66 (1 H, d, J=14 Hz), 4.98(1 H, H, d, J=14 Hz), 5.04(1 H, H, d, J=5 Hz), 5.76(1 H, H, dd, J=8 Hz, 5Hz), 5.84(1 H, d, J=8 Hz), 7.38-7.50 (5 H, m), 7.57 (1 H, d, J=8.5 Hz), 8.16 (1 H, d, J=8.5 Hz), 9.00 (1 H, s), 9.52 (1 H, d, J=8 Hz), 10.09 (1 H, d, J=8 Hz).

Example 8 7ss-[D-2-(7,8-Dlacetoxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3 acetoxymethyl-3-cephem-4-carboxylic Acid a) 7,8-Diacetoxychromone-3-carbonyl Chloride A mixture of 7,8-diacetoxychromone-3-carboxylic acid (9.5 g, 31 mmol), thionyl chloride (2.6 ml), N,N-dimethylformamide (0.1 ml) and benzene (300 ml) was refluxed for 1.5 hours. After the addition of thionyl chloride (2.6 ml) and N,N-dimethylformamide (0.1 ml), the mixture was refluxed for a further hour and evaporated to dryness. The residue was triturated with n-hexane to afford the desired compound (9.3 g. 92.6%).

IR spectrum (cm~', nujol): 1780, 1770, 1670, 1620.

b) 7ss-[D-2-(7,8-Diacetoxychromone-3-carobxamido)-2-(4-hydroxyphehnyl)acetamido]-3 acetoxymethyl-3-cephem-4-carboxylic Acid N,0-Bis(trimethylsilyl)acetamide (1.23 ml) was added dropwise to a suspension of 7p-[D-2- amino-2-(4-hydroxyphenyl)acetamido]-3-acetoxymethyl-3-cephem-4-carboxylic acid (211 mg) in ethyl acetate (5 ml) at OOC. The mixture was stirred for 1 hour at OOC, to which was added the acid chloride (162 mg) described in a).After stirring for 4 hours at OOC, the reaction mixture was diluted with ethyl acetate (150 ml), washed successively with water, 0.5 N hydrochloric acid, water and saturated brine and dried (Mg504). The solvent was evaporated and acetone (10 ml) was added to the residue. The mixture was stirred for 4 hours at room temperature, concentrated to a volume of ca. 5 ml and purified by thin layer chromatography on silica gel to afford the desired compound (24 mg).

Melting point: 200-210 C (decomposition) Elemental analysis: for C32H2,N30,4S C H N Calculated (96): 54.16 3.84 5.92 Found (%): 50.34 3.75 6.34 IR spectrum (cm-1. nujol): 1786, 1775, 1770, 1740, 1710, 1675, 1655, 1630, 1610, 1600, NMR spectrum (#, DMSO-d6): 2.04 (3H,s), 2.39(3H,s), 2.45 (3H,s), 3.48(2H,m), 4.68(1H,d,J=13Hz), 4.96(1H,d, J=13 Hz), 5.04(1 H, H, d, J=5 Hz), 5.72 (1 H, d, J=8 Hz), 5.76 (1 H, m), 6.74(2 H, H, d, J=8.5 Hz), 7.26 (2 H, d, J=8.5 Hz), 7.58 (1 H, d, J=9 Hz), 8.14 (1 H, d, J=9 Hz), 9.00 (1 H, s), 9.40(1 H, H, d, J=8 Hz), 9.46 (1H,s), 9.97(1H,d,J=8Hz).

Example 9 7ss-[D-2-(7,8-Dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3 cephem-4-carboxylic Acid and its Sodium Salt a) 7,8-Dihydroxychromone-3-carbonyl Chloride A mixture of 7,8-dihydroxychromone-3-carboxylic acid (6.6 g, 30 mmol) and thionyl chloride (25 ml) was refluxed for 1 hour, and the thionyl chloride was removed. After the addition of benzene to the residue, the mixture was evaporated to dryness and the residue was triturated with n-hexane to give the desired compound (7.2 g).

IR spectrum (cm~', nujol): 1775, 1660, 1620.

b) 7ss-[D-2-(7,8-Dihydroxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3 cephem4-carboxylic Acid N,O-Bis(trimethylsilyl)acetamide (13.6 ml, 55 mmol) was added dropwise to a stirred suspension of cephaloglycin (4.05 g, 10 mmol) in dichloromethane (100 ml) at OOC. The mixture was stirred for 1 hour at OOC, to which was added the acid chloride (2.3 g, 9.5 mmol). After stirring for 1.5 hours at OOC, the reaction mixture was diluted with ethyl acetate (500 ml), washed successively with saturated brine - 1 N hydrochloric acid (1 200 mlx4) and saturated brine (200 mlx 6) and dried (MgS04).

Removal of the solvent afforded solid, to which was added acetone (100 ml). The mixture was stirred at 250C. overnight and concentrated to a volume of 10 ml. Ethyl acetate and n-hexane were added successively to the residual mixture. The precipitate formed was filtered off and dried to afford the desired compound (1.1 g. 18.8%).

Melting point: 235-2 500C (decomposition) Elemental analysis: for C28H23N30"S C H N Calculated (%): 55.17 3.80 6.89 Found (%): 53.21 3.47 5.45 IR spectrum (cm-t, nujol): 1780, 1740, 1720, 1660, 1615.

NMR spectrum (# DMSO-d6): 2.03(3H,s), 3.44(1H,d,J=18Hz), 3.48(1H,d,J=18Hz), 4.68(1H,d,J=13Hz), 4.98(1H,d,) J=1 3 Hz), 5.04 (1 H, d, J=5 Hz), 5.76 (1 H, m), 5.86 (1 H, d, J=8 Hz), 7.08 (1 H, d, J=8 Hz), 7.28- 7.52 (5 H, m), 7.56(1 H, H, d, J=8 Hz), 8.92(1 H, 5), 9.59(1 H, d, J=8 Hz), 10.34(1 H, H, d, J=8 Hz).

c) Sodium Salt of the Compound Described in b) A suspension of the compound (0.98 g, 1.6 mmol) described in b) in acetone (200 ml) was filtered and, to the filtrate was added sodium 2-ethyl hexanoate (0.5 M solution in ethyl acetate, 3.04 ml) followed by ethyl acetate-ethyl ether (1 :1,200 ml). The precipitate formed was filtered off, washed with ethyl acetate-ethyl ether (1 :1) and dried to afford the desired compound (0.871 g, 85.8%).

Melting point: 230-2400C (decomposition) Elemental analysis: for C28H22N3NaO11S C H N Calculated (%): 53.25 3.67 6.65 Found (%): 49.67 3.29 5.00 IR spectrum (cm-1, nujol): 1760,1660,1610.

NMR spectrum (, DMSO-d6): 2.00(3H,s), 3.19(1H,d,J=18Hz), 3.40(1H,d,J=18Hz), 4.64(1H,d,J=12Hz), 4.98 (2H, m), 5.60 (1 H, m), 5.78 (1 H, d, J=8 Hz), 7.04 (1 H, d, J=8 Hz), 7.12-7.56 (6 H, m), 8.90 (1 H, s), 9.40 (1 H, br.s), 10.42 (1 H, d, J=8 Hz).

The compounds obtained in these examples were tested for their antibacterial activities in vitro.

Example 10 7ss-[D-2-(7,8-Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic Acid and its Sodium Salt a) 7,8-Dihydroxychromone-3-carbonyl Chloride A mixture of 7,8-dihydroxychromone-3-carboxylic acid (6.6 g, 30 mmol) and thionyl chloride (25 ml) was refluxed for 1 hour, and the thionyl chloride was removed. After the addition of benzene to the residue, the mixture was evaporated to dryness and the residue was triturated with n-hexane to give the desired compound (7.2 g).

IR spectrum (cm-1, nujol): 1775, 1660, 1620.

b) 7ss-[D-2-(7,8-Dihydroxychromone-3-carboxamido)-2-(4-hydroxyphanyl0acetamido]-3acetoxymethyl-3-cephem-4-carboxylic Acid N,O-Bis)trimethylsilyl)acetamide (17.3 ml) was added dropwise to a stirred suspension of 2-amino-2-(4-hydroxyphenyl)acetamido]-3-acetoxymethyl-3-cephem-4-cartoxylic acid (4.8 g) in ethyl acetate (200 mi) at 20C. The mixture was stirred for 2 hours at room temperature and cooled to 20C, to which was added the acid chloride (2.4 g) described in a). After stirring for 4 hours at 20C, the reaction mixture was poured into an ice-cooled 0.5 N hydrochloric acid (800 ml) and the mixture was stirred for 30 minutes in an ice bath. The precipitate formed was filtered off, washed with water and dried to afford a crude product (6.5 g).A portion of the product (5.6 g) was stirred in methanol (200 ml) for 2 hours at room temperature and the mixture was filtered. The filtrate was concentrated to a volume of 30 ml and the crystals formed were filtered off, washed successively with methanol and ethyl ether to afford the desired compound (1.2 g).

Melting point: 220-2500C (decomposition) Elemental analysis: for C28H23N3012S C H N Calculated (%): 53.76 3.71 6.72 Found (%): 50.34 3.78 6.34 IR spectrum (cm-1, nujol): 3250, 1770, 1720, 1665, 1615, NMR spectrum (#, DMSO-d6): : 2.04 (3 H, s), 3.48 (2 H, m), 4.68 (1 H, d, J=14 4Hz) 5.00(1 H, H, d, J=14 Hz), 5.04(1 H, H, d, J=5 Hz), 5.76 (2 H, m), 6.75 (2 H, d, J=8 Hz), 7.06 (1 H, d, J=8.5 Hz), 7.28 (2 H, d, J=8 Hz), 7.56 (1 H, d, J=8.5 Hz), 8.94 (1 H, s), 9.38 (1 H, d, J=8 Hz), 10.24(1 H, H, d, J=8 Hz).

c) Sodium Salt of the Compound Described in b) According to the method described in Example 4-c), the desired compound (623 mg) was obtained from the compound (626 mg) described in b).

Melting point: 210-230 C (decomposition) Elemental analysis: for C28H22N3NaO,2S C H N Calculated (%): 51.93 3.42 6.49 Found (%): 49.46 3.88 6.67 IR spectrum (cm-1, nujol): 3250, 1770, 1670, 1620, NMR spectrum (#, DMSO-d6): 2.02 (3H,s), 3.24 (1 H, d, J=18 Hz), 3.44(1 H, d, J=18 Hz), 4.80(1 H, d, J=12 Hz), 4.94(1 H, d, J=4.5 Hz), 5.04(1 H, H, d, J=12 Hz), 5.68 (2 H, m), 6.76 (2 H, d, J=8 Hz), 7.00(1 H, H, d, J=8.5 Hz), 7.28 (2 H, d, J=8 Hz), 7.46 (1 H, d, J=8.5 Hz), 8.88 (1 H, s), 9.02 (4 H, bs), 10.36 (1 H, d, J=8 Hz).

Method Minimal inhibitor concentration (MIC) was determined by the standard agar dilution method of the Japan Society of Chemotherapy.

The compounds described in Examples 1, 2, 3, 4, 9 and 10 were dissolved in sterilized water; the compounds described in Examples 7 and 8 were dissolved in acetone-water (1:1); and the control compounds, cephaloglycin and cephalexin monohydrate, were dissolved in a 3% sodium hydrogen carbonate solution.

Serial two-fold dilutions were made from the above solutions.

One-ml aliquots of each dilution were mixed with 9 ml of Heart influsion agar or Mueller Hinton agar in petri-dishes to make agar plates containing the compound at serially diluted concentrations.

After agar hardened, plates were put in an incubator at 370C for 1.5-2 hours with the lids slightly open to evaporate acetone off the plates.

Test organisms were grown for 18 hours at 370C in Tripticase Soy broth and diluted in saline to approximately 106 colony forming units per ml. A loopful of each celle suspension was applied on the agar plate mentioned above and the plates were incubated for 18 hours at 370C before MIC was determined.

MIC values of the compounds described in Examples 1, 2, 3, 4, 9 and 10 were determined as their sodium salts and the compounds described in Examples 7 and 8 were determined as a free carboxylic acid.

The results are shown in Table 2.

Table 2 MIC (,ug/ml) Test Bacteria Staphylococus Escherichia Klebsiella Proteus Pseudomonas Serratia aureus coli pneumoniae organic aeruginosa marcescens Test Compound 209P NIHJ EK-6 EP-14 EP-172 ES-75 1* 3.13 1.56 < 0.1 50 0.8 0.4 6.25 12.5 0.2 100 3.13 6.25 3* 1.56 0.8 < 0.1 12.5 0.8 0.2 3.13 3.13 0.2 50 3.13 3.13 7* 1.56 0.8 < 0.1 3.13 1.56 0.8 8** 6.25 3.13 0.4 25 3.13 25 9* 1.56 3.13 0.2 12.5 1.56 1.56 10** 0.8 12.5 0.2 100 1.56 100 Cephaloglycin*,** 0.8 3.13 1.56 25 > 100 > 100 Cephalexin* monohydrate 1.56 6.25 3.13 > 100 > 100 > 100 * Heart infusion agar was used ** Mueller Hinton agar was used.

Example 11 A vial was aseptically sealed, so as to involve 125 mg of sodium salt of 7p-[D-2-(6,7diacetoxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3-cephem-4-carboxylic acid. In using, 5 ml of distilled water for injection was added thereto, so that injection may be obtained.

Example 12 Tablet Sodium salt of 7P-[D-2-(6,7-dihydroxychromone-3-carboxa mido)2 phenylaceta mido]-3-acetoxymethyl-3-cephem-4-carboxylic acid 250 mg Crystalline cellulose 80 mg Calcium salt of carboxymethyl cellulose 38 mg Calcium stearate 2 mg One Tablet 370 mg Tablet was prepared with the above formulation by means of conventional manner.

Example 13 Formulation for tablet 7ss-[D-2-(6,7Dihydroxychromone-3-carboxa mido)-2-(4 hydroxyphenyl)aceta mido]-3-acetoxymethyl-3-cephem-4-carboxylic acid 250 mg Crystalline cellulose 80 mg Calcium salt of carboxymethylcellulose 38 mg Calcium stearate 2 mg 1 Tablet 370 mg The table was produced by usual method with the above formulation.

Claims

1. A cephalosporin derivative represented by the general formula:

wherein R', R2 and R3 each represents hydroxy, acyloxy, and hydrogen provided that two or three of R1, R2 and R3 are not hydrogen; and R4 represents hydrogen or hydroxy; or a non-toxic salt thereof.

2.7ss-[D-2-(6,7-diacetoxychromone-3-carboxamido)-2-phenylacetamido]-3-acetoxymethyl-3 cephem-4-carboxylic acid and salts thereof.

3.7ss-[D-2-(6,7-diacetoxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic acid and salts thereof.

4.7,B-[D-2-(6,7-dihydroxychromone-3-carboxamido)-2-phenylaceta mido]-3-acetoxymethyl-3- cephem-4-carboxylic acid and salts thereof.

5.7ss-[D-2-(6,7-dihydroxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic acid and salts thereof.

6. 7-[D-2-(7,8-diacetoxychrnmone-3-carboxa mido)-2-phenylacetamido]-3-acetoxymethyl-3cephem-4-carboxylic acid and salts thereof.

7,7ss-[D-2-(7,8-diacetoxychromone-3-carboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic acid and salts thereof.

8.7ss-[D-2-(7,8-dihydroxychromone-3-carobxamido)-2-phenylacetamido]-3cephem-4-carboxylic acid and salts thereof.

9.7p-[D-2-dihydroxychromone-3-ca rboxamido)-2-(4-hydroxyphenyl)acetamido]-3acetoxymethyl-3-cephem-4-carboxylic acid and salts thereof.

10. A process for the preparation of a cephalosporin derivative represented by the general formula (I) as defined in Claim 1, which comprises reacting a compound represented by the general formula (all):

wherein R4 represents hydrogen or hydroxy: or a salt or hydrate thereof, with a compound represented by the general formula (III):

wherein R1, R2 and R3 are as defined, or with a reactive derivative thereof.

11. A process for the production of a compound having the general formula (1)-i:

wherein R', R2 and R3 each represents hydroxy or hydrogen provided that two or three of R', R' and R3 are not hydrogen at the same time, and R4 represents hydrogen or hydroxy; or its non-toxic salts, characterized by hydrolyzing a compound represented by the general formula (1)-2:

wherein R", R" and R "each represents acyloxy or hydrogen provided that two or three of R",R" and R " are not hydrogen at the same time, and R4 is as defined.

12. A pharmaceutical composition which comprises a cephalosporin derivative of the general formula (I) as defined in any of claims 1 to 9, together with a pharmaceuticaily acceptable carrier or diluent.