Enantioselective oligomerization of alpha-hydroxy carboxylic acids and alpha-amino acids

Abstract

An enzymatic synthesis and composition of oligomers and co-oligomers comprised of α-hydroxy carboxylic acids and α-amino acids or peptides is disclosed. In a preferred embodiment, a α-hydroxy carboxylic acid with a specific chiral configuration is linked by an amide linkage to a α-amino acid specific with a specific chiral configuration or linked by an amide linkage to a peptide made up of α-amino acid monomers having identical chiral configurations. Proteolytic enzymes catalyze oligomerization of the α-hydroxy carboxylic acid and α-amino acid. The degree and distribution of oligomerization varies upon the type and concentrations of different reaction mixtures utilized and upon the length of allowed reaction time. The resultant oligomers may be provided to animals such as ruminants as bioavailable amino acid supplements that are resistant to degradation in the rumen and other animals such as swine, poultry and aquatic animals.

1 . A process for the preparation of an oligomer, the process comprising: 
 forming a reaction mixture comprising an enzyme, and an enantiomeric mixture of an α-amino acid or a derivative thereof; and    forming an oligomer from the reaction mixture, said oligomer incorporating one of the members of the enantiomeric mixture of the α-amino acid or derivative thereof in preference to the other member.    
   
   
       2 . A process as set forth in  claim 1 , wherein the enantiomeric mixture is a racemic mixture.  
   
   
       3 . A process as set forth in  claim 1 , wherein the oligomer comprises an α-amino acid or a derivative thereof comprising an oligomer of two or more α-amino acid monomers.  
   
   
       4 . A process as set forth in  claim 3 , wherein the α-amino acid monomers comprise L-α-amino acid monomers.  
   
   
       5 . A process as set forth in  claim 1 , wherein the reaction mixture further comprises an α-hydroxy carboxylic acid or a derivative thereof and the formed oligomer comprises an oligomer of two or more α-amino acid monomers bonded to a residue of the α-hydroxy carboxylic acid or a derivative thereof by an amide or ester linkage.  
   
   
       6 . A process as set forth in  claim 5  wherein said α-hydroxy carboxylic acid or a derivative thereof is present in said reaction mixture in an enantiomeric mixture.  
   
   
       7 . A process as set forth in  claim 6  wherein one α-hydroxy carboxylic acid enantiomer of the enantiomeric mixture is bonded to the oligomer in preference to the other enantiomer of said enantiomeric mixture.  
   
   
       8 . A process as set forth in  claim 1  wherein the α-amino acid is selected from the group consisting of methionine, lysine, or a derivative thereof.  
   
   
       9 . A process as set forth in  claim 1  wherein said enantiomeric mixture of α-amino acids comprises methionine and lysine.  
   
   
       10 . A process as set forth in  claim 1  wherein the enzyme comprises a protease.  
   
   
       11 . A process as set forth in  claim 10  wherein the enzyme comprises a protease selected from the group consisting of papain, bromelain, cathepsin s, cathepsin b, capthesin c, and substilisin.  
   
   
       12 . A process as set forth in  claim 1  wherein the reaction mixture comprises an aqueous solution.  
   
   
       13 . A process as set forth in  claim 1  wherein the reaction mixture comprises an aqueous phase and an organic solvent phase.  
   
   
       14 . A process as set forth in  claim 1  wherein the reaction mixture comprises an α-hydroxy carboxylic acid derivative comprising a free acid, an acid halide, an amide, an ester or an anhydride.  
   
   
       15 . A process as set forth in  claim 1  wherein the reaction mixture comprises an α-amino acid derivative comprising a free acid, an acid halide, an amide, an ester or an anhydride.  
   
   
       16 . A composition comprising a residue of an α-hydroxy carboxylic acid bonded to a peptide by an amide linkage, said peptide comprising two or more α-amino acid residues, each of said α-amino acids being independently selected from the group consisting of α-amino acids.  
   
   
       17 . A composition as set forth in  claim 16  wherein more than 50% of the α-amino acid residues in the peptide are of identical chirality.  
   
   
       18 . A composition as set forth in  claim 17  wherein essentially all of the α-amino acid residues in the peptide are of identical chirality.  
   
   
       19 . A composition comprising a residue of an α-hydroxy carboxylic acid bonded to a peptide by an ester linkage, said peptide comprising two or more α-amino acid residues, each of said α-amino acids being independently selected from the group consisting of α-amino acids.  
   
   
       20 . A composition as set forth in  claim 19  wherein more than 50% of the α-amino acid residues in the peptide are of identical chirality.  
   
   
       21 . A composition as set forth in  claim 20  wherein essentially all of the α-amino acid residues in the peptide are of identical chirality.  
   
   
       22 . A composition comprising:  
     
       
         
         
             
             
         
       
     
     wherein: 
 R 1  is hydrogen, hydrocarbyl or substituted hydrocarbyl,  
 R 2  is hydrogen, hydrocarbyl, substituted hydrocarbyl, or a hydroxy protecting group,  
 R 3  is hydrogen, hydrocarbyl or substituted hydrocarbyl,  
 each AA is the residue of an α-amino acid or derivative thereof wherein greater than one-half of the AA residues are derived from α-amino acids or derivatives thereof having the an identical chiral configuration, and  
 n is at least 2.  
 
   
   
       23 . The composition of  claim 22  wherein R 1  is CH 3 SCH 2 CH 2 —.  
   
   
       24 . The composition of  claim 23  wherein R 2  is H.  
   
   
       25 . The composition of  claim 22  wherein R 2  is H.  
   
   
       26 . The composition of  claim 22  wherein n is at least 4 and no more than 12 and each AA is methionine.  
   
   
       27 . The composition of  claim 22  wherein n is at least 3 and no more than 5 and each AA is selected from the group consisting of methionine and lysine.  
   
   
       28 . The composition of  claim 22  wherein n is at least 2 and no more than 10 and each AA is selected from the group consisting of methionine and lysine.  
   
   
       29 . A process of providing a food ration to an animal comprising providing the composition of  claim 22  to the animal wherein the method of administration is selected from the group consisting of oral administration, eye spray, placement in ear, placement in nasal cavity, bucchal administration, sublingual administration, rectal administration and injection.  
   
   
       30 . A process as set forth in  claim 29  wherein the composition is orally administered to the animals.  
   
   
       31 . A process as set forth in  claim 30  wherein the animal is selected from the group consisting of ruminants, swine, poultry, and aquatic animals.  
   
   
       32 . A process as set forth in  claim 31  wherein the ruminant is a dairy cow or beef cattle.  
   
   
       33 . A process as set forth in  claim 32  wherein the cow is a lactating dairy cow.  
   
   
       34 . A process as set forth in  claim 31  wherein the aquatic animal is a fish or crustacean.  
   
   
       35 . A process as set forth in  claim 34  wherein the fish is selected from the group consisting of freshwater and salt water fish.  
   
   
       36 . A process as set forth in  claim 35  wherein the freshwater and salt water fish are selected from the group consisting of carp, trout, catfish, bass, sea bass, cod, and salmon.  
   
   
       37 . A process as set forth in  claim 34  wherein the crustaceans are selected from the group consisting of shrimp, crabs, lobster, and freshwater shrimp.  
   
   
       38 . A process for providing a food ration to an animal, the process comprising: providing an oligomer or co-oligomer composition prepared from a mixture containing an enzyme, an α-amino acid or a derivative thereof, and optionally, an α-hydroxy carboxylic acid or a derivative thereof, 
 wherein the method of administration is selected from the group consisting of oral administration, eye spray, placement in ear, placement in nasal cavity, bucchal administration, sublingual administration, rectal administration and injection.    
   
   
       39 . A process as set forth in  claim 38  wherein the animal is selected from the group consisting of ruminants, swine, poultry and aquatic animals.  
   
   
       40 . A process as set forth in  claim 39  wherein the ruminant is a dairy cow or beef cattle.  
   
   
       41 . A process as set forth in  claim 40  wherein the cow is a lactating dairy cow.  
   
   
       42 . A process as set forth in  claim 39  wherein the aquatic animal is a fish or crustacean.  
   
   
       43 . A process as set forth in  claim 42  wherein the fish is selected from the group consisting of freshwater and salt water fish.  
   
   
       44 . A process as set forth in  claim 43  wherein the freshwater and salt water fish are selected from the group consisting of carp, trout, catfish, bass, sea bass, cod, and salmon.  
   
   
       45 . A process as set forth in  claim 42  wherein the crustaceans are selected from the group consisting of shrimp, crabs, lobster, and freshwater shrimp.  
   
   
       46 . An orally administered dietary supplement comprising a vitamin, a mineral or a nutrient coated with an oligomeric coating, said coating comprising a residue of an α-hydroxy carboxylic acid bonded to a peptide by an amide linkage, said peptide comprising two or more α-amino acid residues, each of said α-amino acids being independently selected from the group consisting of α-amino acids.  
   
   
       47 . A process for providing an animal with a dietary supplement comprising a vitamin, mineral or nutrient, the process comprising: 
 coating said vitamin, mineral or nutrient with a composition to form a dietary supplement, said composition comprising a residue of an α-hydroxy carboxylic acid bonded to a peptide by an amide linkage, said peptide comprising two or more α-amino acid residues, each of said α-amino acids being independently selected from the group consisting of α-amino acids; and    orally administering the dietary supplement to the animal.    
   
   
       48 . A process as set forth in  claim 47  wherein the peptide comprises methionine.  
   
   
       49 . A process as set forth in  claim 47  wherein the animal is selected from the group consisting of ruminants, swine, poultry, and aquatic animals.  
   
   
       50 . A process as set forth in  claim 49  wherein the ruminant comprises a dairy cow or beef cattle.  
   
   
       51 . A process as set forth in  claim 50  wherein the ruminant comprises a lactating dairy cow.  
   
   
       52 . A process as set forth in  claim 49  wherein the aquatic animal comprises a fish or crustacean.  
   
   
       53 . A process as set forth in  claim 52  wherein the fish is selected from the group consisting of freshwater and salt water fish.  
   
   
       54 . A process as set forth in  claim 53  wherein the freshwater and salt water fish are selected from the group consisting of carp, trout, catfish, bass, sea bass, cod, and salmon.  
   
   
       55 . A process as set forth in  claim 52  wherein the crustaceans are selected from the group consisting of shrimp, crabs, lobster, and freshwater shrimp.  
   
   
       56 . A process for providing an animal with a dietary supplement comprising a vitamin, mineral or nutrient, the process comprising: 
 preparing a mixture comprising an enzyme and at least one α-amino acid, each α-amino acid being present in the mixture as a free acid, acid halide, amide, ester or anhydride independently of the other,    forming an α-amino acid oligomer in the mixture,    coating said vitamin, mineral or nutrient with the α-amino acid oligomer to form an α-amino acid oligomer coated dietary supplement, and    orally administering the dietary supplement to the animal.    
   
   
       57 . A process as set forth in  claim 56  wherein the animal comprises an aquatic animal selected from the group consisting of shrimp, carp, and trout.  
   
   
       58 . A process as set forth in  claim 57 , wherein the α-amino acid oligomeric coating comprises methionine.  
   
   
       59 . A process for purifying an α-hydroxy carboxylic acid enantiomer or derivative thereof from an enantiomeric mixture, the process comprising: 
 forming a reaction mixture comprising (i) an enantioselective enzyme, (ii) an enantiomeric mixture of an α-hydroxy carboxylic acid or a derivative thereof, and (iii) an enantiomeric mixture of an α-amino acid or a derivative thereof;    forming a reaction product from the reaction mixture, the reaction product comprising (i) an oligomer having a first α-hydroxy carboxylic acid enantiomer of the enantiomeric mixture incorporated therein in preference to the second enantiomer, and (ii) unreacted second enantiomer; and    separating the oligomer and unreacted second enantiomer from the reaction product and each other.    
   
   
       60 . A process as set forth in  claim 59 , wherein the enantioselective enzyme promotes an esterification reaction.  
   
   
       61 . A process as set forth in  claim 60 , wherein the enantioselective enzyme is a lipase enzyme.  
   
   
       62 . A process as set forth in  claim 59 , wherein the first enantiomer is recovered from the separated oligomer by hydrolyzing the oligomer with acid and separating the first α-hydroxy carboxylic acid enantiomer or derivative thereof from other hydrolyzates.  
   
   
       63 . A process as set forth in  claim 62 , wherein the first α-hydroxy carboxylic acid enantiomer or derivative thereof is separated by subjection to enantioselective chromatography.  
   
   
       64 . A process as set forth in  claim 59 , wherein the second unreacted α-hydroxy carboxylic acid enantiomer or derivative thereof is recovered from the reaction mixture by rotary evaporation.  
   
   
       65 . A process as set forth in  claim 59 , wherein the α-hydroxy carboxylic acid comprises 2-hydroxy-4-(methylthio)butyric acid or a derivative thereof.  
   
   
       66 . A process as set forth in  claim 59  wherein the process further comprises recovering the first enantiomer from said separated oligomer by by hydrolyzing the oligomer with acid and separating the first α-hydroxy carboxylic acid enantiomer or derivative thereof from other hydrolyzates; and 
 converting a fraction of the recovered first enantiomer to the stereochemical form of the second enantiomer thereby forming an enantiomeric mixture comprising the first and second enantiomers.    
   
   
       67 . A process as set forth in  claim 59  wherein the process further comprises recovering the unreacted second enantiomer from the reaction product; and 
 converting a fraction of the separated unreacted second enantiomer to the stereochemical form of the first enantiomer thereby forming an enantiomeric mixture comprising the first and second enantiomers, at least a portion of the first enantiomer in the enantiomeric mixture being derived from separated unreacted second enantiomer.    
   
   
       68 . A process as set forth in  claim 67  wherein the enantiomeric mixture formed from the recovered second enantiomer is recycled for re-use in the formation of a reaction mixture.  
   
   
       69 . A process as set forth in  claim 67 , wherein the enzyme is removed from the reaction mixture and recycled.  
   
   
       70 . A process as set forth in  claim 69 , wherein the enzyme is removed from the reaction mixture by size exclusion chromatography.  
   
   
       71 . A process as set forth in  claim 67  wherein the second unreacted α-amino acid enantiomers or derivatives thereof are separated from the reaction mixture by rotary evaporation.  
   
   
       72 . A process for purifying an enantiomeric mixture of α-amino acid or derivative thereof, the process comprising: 
 forming a reaction mixture comprising (i) an enzyme, (ii) an enantiomeric mixture of α-amino acid or a derivative thereof, and (iii) an α-hydroxy carboxylic acid or a derivative thereof;    forming a reaction product from the reaction mixture, the reaction product comprising (i) an oligomer incorporating a first member of the enantiomeric mixture of α-amino acid or derivative thereof in preference to a second enantiomer of the enantiomeric mixture or derivative thereof, and (ii) unreacted second enantiomer; and    separating the oligomer and unreacted second enantiomer from the reaction product and each other.    
   
   
       73 . A process as set forth in  claim 72 , wherein the first enantiomer is recovered from the separated oligomer by hydrolyzing the oligomer with acid and separating the α-amino acid enantiomer or derivative thereof from other hydrolyzates.  
   
   
       74 . A process as set forth in  claim 73 , wherein the α-amino acid enantiomer or derivative thereof is separated by subjection to enantioselective chromatography.  
   
   
       75 . A process as set forth in  claim 72 , wherein the unreacted α-amino acid enantiomer or derivative thereof is recovered from the reaction mixture by rotary evaporation.  
   
   
       76 . A process as set forth in  claim 72 , wherein the α-amino acid is selected from the group consisting of methionine and lysine.  
   
   
       77 . A process as set forth in  claim 72 , wherein the α-hydroxy carboxylic acid comprises 2-hydroxy-4-(methylthio)butyric acid or a derivative thereof.  
   
   
       78 . A process for purifying an α-amino acid enantiomer or derivative thereof in an enantiomeric mixture, the process comprising: 
 forming a reaction mixture comprising (i) an enantioselective enzyme and (ii) an enantiomeric mixture of an α-amino acid or derivative thereof;    forming a peptide reaction product mixture comprising (i) an oligomer or a co-oligomer having a first enantiomer of the enantiomeric mixture incorporated therein in preference to the second enantiomer of the enantiomeric mixture, and (ii) unreacted second enantiomer; and    separating the oligomer or co-oligomer and unreacted second enantiomer from the reaction product mixture and each other    
   
   
       79 . A process as set forth in  claim 78 , wherein the first enantiomer is recovered from the separated oligomer or co-oligomer by hydrolyzing the oligomer or co-oligomer with acid and separating the α-amino acid enantiomer or derivative thereof from other hydrolyzates.  
   
   
       80 . A process as set forth in  claim 79 , wherein the first enantiomer or derivative thereof is separated by subjection to enantioselective chromatography.  
   
   
       81 . A process as set forth in  claim 78 , wherein the unreacted α-amino acid enantiomer or derivative thereof is recovered from the reaction mixture by rotary evaporation.  
   
   
       82 . A process as set forth in  claim 78  wherein the α-hydroxy carboxylic acid derivative is a free acid, acid halide, amide, ester or anhydride.  
   
   
       83 . A process as set forth in  claim 82  wherein the α-amino acid derivative is a free acid, acid halide, amide, ester or anhydride.