Process for preparing cephalosporanic acid derivatives using alpha-ketoacid derivatives

Abstract

A process for preparing cephalosporanic acid derivatives comprises the steps of enzymatically converting a 3-thiolated cephalosporin C compound of formula III:— into a 3-thiolated-α-ketoadipyl-7-aminocephalosporanic acid derivative of formula IV: wherein R is a heterocyclic group comprising at least a nitrogen atom. Compounds of formula IV are used in the preparation of cephalosporin C antibiotics and derivatives thereof.

1 . A process for preparing cephalosporanic acid derivatives comprising the steps of:—
 enzymatically converting a 3-thiolated cephalosporin C compound of formula III:— 
                     
  into a 3-thiolated-α-ketoadipyl-7-aminocephalosporanic acid derivative of formula IV:  
                     
  wherein r is a heterocyclic group comprising at least a nitrogen atom.  
 
     
     
         2 . A process as claimed in  claim 1  wherein the compound of formula III is enzymatically converted into a compound of formula IV by an immobilised enzyme system.  
     
     
         3 . A process as claimed in  claim 2  wherein the enzyme system comprises co-immobilised D-Amino acid oxidase and catalase.  
     
     
         4 . A process as claimed in  claim 3  wherein the enzymatic conversion is carried out in the presence of molecular oxygen, at a pressure of 1 to 5 bar absolute, a pH of from 6.5 to 8.0 and at a temperature of from 15 to 30° C. for a period of from 30 mins to 180 mins.  
     
     
         5 . A process as claimed in  claim 1  comprising the step of separating the enzyme system from the reaction mixture, preferably by filtration.  
     
     
         6 . A process as claimed in  claim 1  including the step of purifying the compound of formula IV.  
     
     
         7 . A process as claimed in  claim 6  wherein the compound is purified using an adsorption column.  
     
     
         8 . A process as claimed in  claim 1  wherein the enzymes are co-immobilised using a suitable cross-linker agent in a suitable solid support.  
     
     
         9 . A process as claimed in  claim 8  wherein the enzymes are in the form of crystals of a size suitable for use as a biocatalyst.  
     
     
         10 . A process as claimed in  claim 1  wherein the enzymatic processes are carried out while maintaining the enzyme in dispersion in an aqueous substrate solution.  
     
     
         11 . A process as claimed in  claim 1  wherein the or each enzymatic process is carried out in a column.  
     
     
         12 . A process as claimed in  claim 1  including the step of recovering the enzyme for reuse.  
     
     
         13 . A process as claimed in  claim 1  wherein the compound of formula IV is used without purification in a continuous process for obtaining any useful derivative.  
     
     
         14 . A process as claimed in  claim 1  wherein R is a heterocyclic group comprising at least one nitrogen atom and optionally a sulphur or oxygen atom.  
     
     
         15 . A process as claimed in  claim 14  wherein R is a heterocyclic group selected from any one or more of the group comprising thienyl, diazolyl, tetrazolyl, thiazolyl, triazinyl, oxazolyl, oxadiazolyl, pyridyl, pirimidinyl, benzo thiazolyl, benzimidazolyl, benzoxazolyl, or any derivative thereof, preferably 5-methyl-1,3,4-thiadiazol-2-yl, 1-methyl-1H-tetrazol-5-yl or 1,2,5,6-tetrahydro-2-methyl-5,6-dioxo-1,2,4-triazin-3-yl.  
     
     
         16 . A 3-thiolated-α-ketoadipyl-7-aminocephalosporanic acid derivative of formula IV whenever prepared by a process as claimed in  claim 1 .  
     
     
         17 . A compound of the Formula:— 
       
         
           
           
               
               
           
         
          wherein in formula IV, R is 1,2,5,6-tetrahydro-2-methyl-5,6-dioxo-1,2,4-triazin-3-yl.  
       
     
     
         18 . A compound of the Formula:— 
       
         
           
           
               
               
           
         
          wherein in formula IV, R is 1-methyl-1H-tetrazol-5-yl.  
       
     
     
         19 . Use of a compound of formula IV as defined in  claim 1  as an intermediate in a process for preparing cephalosporin C antibiotics.  
     
     
         20 . Use of an intermediate compound of the formula:— 
       
         
           
           
               
               
           
         
       
       in a process for preparing cephalosporin C antibiotics wherein in formula IV R is 5-methyl-1,3,4-thiadiazol-2-yl.  
     
     
         21 . A process for preparing cephalosporanic acid derivatives as claimed in  claim 1  comprising the step of: 
 enzymatically converting a compound of formula IV to form a compound of formula I  
                     
  wherein R is a heterocyclic group comprising at least one nitrogen atom and R 1  and R 2  are both hydrogen atoms or one of them is a hydrogen atom and the other is an acyl donor.  
 
     
     
         22 . A process as claimed in  claim 21  wherein a compound of formula IV is enzymatically converted to form a compound of formula I using Glutaryl-7-ACA acylase.  
     
     
         23 . A process as claimed in  claim 21  wherein the enzymation takes place at a temperature of approximately 20° C. and at a pH of between 6.5 and 8.0.  
     
     
         24 . A process as claimed in  claim 21  wherein the enzyme is imrnmobilised using a suitable cross-linker agent in a suitable solid support.  
     
     
         25 . A process as claimed in  claim 24  wherein the enzyme is in the form of crystals of a size suitable for use as a biocatalyst.  
     
     
         26 . A process as claimed in  claim 21  wherein enzymation is carried out while maintaining the enzyme in dispersion in an aqueous substrate solution.  
     
     
         27 . A process as claimed in  claim 21  wherein the enzymatic process is carried out in a column.  
     
     
         28 . A process as claimed in  claim 21  including the step of recovering the enzyme for reuse.  
     
     
         29 . Use of a compound of formula I as defined in  claim 21  as an intermediate in a process for preparing cephalosporin C derivatives.  
     
     
         30 . A process for preparing 3-thiolated cephalosporanic acid derivatives comprising the steps of;—
 enzymatically converting a compound of formula III  
                     
  into a 3-thiolated-α-ketoadipyl-7-aminocephalosporanic acid derivative of formula IV:  
                     
  and enzymatically converting a compound of formula IV to form a 3-thiolated 7-ACA compound of formula I  
                     
  wherein R is a heterocyclic group comprising at least one nitrogen atom and R 1  and R 2  are both hydrogen atoms or one of them is a hydrogen atom and the other is an acyl donor.  
 
     
     
         31 . A process as claimed in  claim 30  wherein the compound of formula III is enzymatically converted into a compound of formula I in one step by an immobilised enzyme system.  
     
     
         32 . A process as claimed in  claim 31  wherein the enzyme system comprises a combination of co-immobilised D-amino acid oxidase/catalase in the presence of immobilised Glutaryl-7-ACA acylase.  
     
     
         33 . A process as claimed in  claim 30  wherein the enzymation takes place at a temperature of approximately 20° C. and at a pH of between 6.5 and 8.0.  
     
     
         34 . A process as claimed in  claim 30  wherein the enzymes are co-immobilised using a suitable cross-linker agent in a suitable solid support.  
     
     
         35 . A process as claimed in  claim 34  wherein the enzymes are in the form of crystals of a size suitable for use as a biocatalyst.  
     
     
         36 . A process as claimed in  claim 30  wherein the enzymatic processes are carried out while maintaining the enzyme in dispersion in an aqueous substrate solution.  
     
     
         37 . A process as claimed in  claim 30  wherein the or each enzymatic process is carried out in a column.  
     
     
         38 . A process as claimed in  claim 30  including the step of recovering the enzyme for reuse.  
     
     
         39 . A process as claimed in  claim 30  wherein the compound of formula III is used without purification in a continuous process for obtaining any useful derivative.  
     
     
         40 . A process for preparing cephalosporanic acid derivatives comprising the steps of:—
 reacting cephalosporin C with a thiol compound of the general formula II  
 R—SH  II  
  wherein R is a heterocyclic group comprising at least one nitrogen atom,  
 to form a 3-thiolated cephalosporin Compound of formula III  
                     
  wherein R is as defined above,  
 and, after formation of the compound of formula III removing excess thiol of formula II.  
 
     
     
         41 . A process as claimed in  claim 40  wherein the excess thiol is removed by adsorption on an anion exchange resin.  
     
     
         42 . A process as claimed in  claim 41  wherein the anion exchange resin is a microporous resin having a cross-linked acrylic copolymer structure.  
     
     
         43 . A process as claimed in  claim 42  wherein the anion exchange resin comprises an 8% cross-linking containing functional thialkyl benzyl ammonium group.  
     
     
         44 . A process as claimed in  claim 41  wherein the resin is in the chloride, hydroxy, phosphate or acetate cycle.  
     
     
         45 . A process as claimed in  claim 40  wherein the excess thiol is removed by crystallisation.  
     
     
         46 . A process as claimed in  claim 45  wherein crystallisation is carried out at an acidic pH.  
     
     
         47 . A process as claimed in  claim 40  wherein the excess thiol is removed by crystallisation followed by adsorption on an anion exchange resin.  
     
     
         48 . A process as claimed in  claim 40  wherein the cephalosporin C is in an aqueous medium.  
     
     
         49 . A process as claimed in  claim 40  wherein the cephalosporin C is in the form of a concentrated cephalosporin C solution.  
     
     
         50 . A process as claimed in  claim 40  wherein the reaction is carried out at a pH of between 5.5 and 8.0, at a temperature of from 60° C. to 80° C., for a period of from 1 to 8 hours.  
     
     
         51 . A process as claimed in  claim 40  wherein the reaction is carried out at a pH of approximately 6.0 and at a temperature of approximately 65° C.  
     
     
         52 . A process as claimed in  claim 40  wherein the thiol compound is present in an amount of between 1 and 5 mol/mol of cephalosporin C.  
     
     
         53 . A process as claimed in  claim 40  wherein R is a heterocyclic group comprising at least one nitrogen atom and optionally a sulphur or oxygen atom.  
     
     
         54 . A process as claimed in  claim 40  wherein R is a heterocyclic group selected from any one or more of thienyl, diazolyl, thiazolyl, tetrazolyl, thiadiazolyl, triazinyl, oxazolyl, oxadiazolyl, pyridyl, pirimidinyl, benzothiazolyl, benzimidazolyl, benzoxazolyl, or any derivative thereof, preferably 5-methyl-1,3,4-thiadiazol-2-yl, 1-methyl-tetrazol-5-yl or 1,2,5,6-tetrahydro-2-methyl-5,6-dioxo-1,2,4-triazin-3-yl.  
     
     
         55 . A compound of formula III  
       
         
           
           
               
               
           
         
         wherein R is a heterocyclic group comprising at least one nitrogen atom,  
         obtained by a process as claimed in any of  claims 40  to  54 .  
       
     
     
         56 . A compound of the Formula:— 
       
         
           
           
               
               
           
         
          wherein in formula III R is 5-methyl-1,3,4-thiadiazol-2-yl.  
       
     
     
         57 . A compound of the Formula:— 
       
         
           
           
               
               
           
         
          wherein in formula III R is 1,2,5,6-tetrahydro-2-methyl-5,6-dioxo-1,2,4-triazin-3-yl.  
       
     
     
         58 . Use of a compound of formula III as defined in  claim 55  as an intermediate in a process for preparing cephalosporin C derivatives.  
     
     
         59 . A process for preparing cephalosporanic acid derivatives comprising the steps of:—
 enzymatically converting a 3-thiolated cephalosporin C compound of formula III obtained by a process as claimed in any of  claims 40  to  54 :— 
                     
  into a 3-thiolated-α-ketoadipyl-7-aminocephalosporanic acid derivative of formula IV:  
                     
  wherein R is a heterocyclic group comprising at least a nitrogen atom.  
 
     
     
         60 . A process as claimed in  claim 59  comprising the step of: 
 enzymatically converting a 3-thiolated α-ketoadipyl 7-ACA compound of formula IV  
                     
  to form a 3-thiolated 7-ACA compound of formula I  
                     
  wherein R is a heterocyclic group comprising at least one nitrogen atom and R 1  and R 2  are both hydrogen atoms or one of them is a hydrogen atom and the other is an acyl donor.  
 
     
     
         61 . A process for preparing cephalosporanic acid dervatives comprising the step of: 
  enzymatically converting a compound of formula IV                           to form a compound of formula I                           wherein R is a heterocyclic group comprising at least one nitrogen atom and R 1  and R 2  are both hydrogen atoms or one of them is a hydrogen atom and the other is an acyl donor.    
     
     
         62 . A process as claimed in  claim 61  wherein a compound of formula IV is enzymatically converted to form a compound of formula I with Glutaryl-7-ACA acylase.