Enzymatic method for the synthesis and separation of peptides

Claim 3

The method recited in claim 1 wherein the uncharged peptide is converted to a charged species by cleavage with an esterase enzyme.

Claim 4

The method recited in claim 1 wherein the condensation enzyme is a peptide forming acidic or neutral protease active in a pH range of about 4.0 to 8.0.

Claim 5

The method recited in claim 1 wherein the condensation enzyme is a protease selected from the group consisting of serine proteinases, thiol proteinases, carboxyl proteinases and metalloproteinases.

Claim 6

The method recited in claim 1 wherein the condensation enzyme is selected from the group consisting of chymotrypsin, trypsin, substilisin BNP', Achromobacter protease, papain, pepsin, thermolysin and prolisin; and the pH of the reaction mixture is between about 4.0 to 8.0.

Claim 7

The method recited in claim 1 wherein the condensation enzyme is thermolysin and the pH of the reaction mixture is about 5.5.

Claim 8

The method recited in claim 5 wherein the membrane comprises a water-immiscible organic liquid immobilized by capillarity in pores in a microporous sheet the organic liquid being a solvent for the uncharged peptide.

Claim 9

The method recited in claim 7 wherein the microporous sheet is made from a material selected from a group consisting of polytetrafluoroethylene and, polypropylene and the solvent comprises at least one compound selected from a group consisting of n-1-decanol, n-hexadecanol, n-dodecanol and mixtures thereof.

Claim 10

The method recited in claim 7 wherein the membrane is a Bend, Type 1 Hollow Fiber Selective Dialysis Membrane.

Claim 11

The method recited in claim 4 wherein the pH of the aqueous solution is between about 4.0 and 6.0.

Claim 12

The method recited in claim 5 wherein the pH of the aqueous solution is between about pH 4.0 and 6.0.

Claim 13

The method recited in claim 6 wherein the first compound is D,L-phenylalanine methyl ester, and the second compound is N-acyl-β-substituted-L-aspartic acid having an α-carboxylate group.

Claim 14

The process recited in claim 12 wherein the N-acyl group is selected from the group consisting of φCH 2 OCO--, --CH═O and φCH 2 CO--; and the β-substituent is selected from the group consisting of φCH 2 O--, ter BuO--, CH 3 O--,NH 2 --.

Claim 15

An enzymatic method for the continuous synthesis of N-acyl-beta-substituted- L- aspartyl- L-phenylalanine, comprising the steps of: reacting a N- acyl- beta-substituted- L-aspartic acid first reactant having a free alpha-carboxylate group with a phenylalanine lower alkyl ester second reactant having an alpha-ammonium group, in an aqueous reaction mixture including a condensation enzyme, to form an uncharged N- acyl-beta- substituted- L- aspartyl- L- phenylalanine lower alkyl ester product under conditions that maintain the reactants in a charged state and the product in an uncharged state; diffusing the uncharged product in the aqueous reaction mixture across an ion rejection membrane into a product mixture by osmosis; and reacting the uncharged product in the product mixture to form a charged N- acyl- beta-substituted- L- aspartyl- L- phenylalanine product that does not back-diffuse into the aqueous reaction mixture so that the chemical equilibrium is pulled toward the production of uncharged product and said diffusing step is enhanced.

Claim 16

An enzymatic method for the continuous synthesis of N-acyl-beta-substituted- L- aspartyl- L-phenylalanine, comprising the steps of: reacting a N- acyl- beta-substituted- L-aspartic acid first reactant having a free alpha-carboxylate group with a phenylalanine lower alkyl ester second reactant having an alpha-ammonium group, in an aqueous reaction mixture including a condensation enzyme, to form an uncharged N- acyl-beta- substituted- L- aspartyl- L- phenylalanine lower alkyl ester product under conditions that maintain the reactants in a charged state and the product in an uncharged state; diffusing the uncharged product in the aqueous reaction mixture across an ion rejection membrane into a product mixture by osmosis; and reacting the uncharged product in the product mixture to form a charged N- acyl- beta-substituted- L- aspartyl- L- phenylalanine product that does not back-diffuse into the aqueous reaction mixture so that the chemical equilibrium is pulled toward the production of uncharged product and said diffusing step is enhanced.