Enzymatic process for the determination of beta-lactam...

Details Enzymatic process for the determination of beta-lactam... Enzymatic process for the determination of beta-lactam...

C - Chemistry, Metallurgy

12

Q

150/15.3, 150/3.

C12Q 1/37 (2006.01) C07K 5/068 (2006.01) A61K 38/00 (2006.01)

Patent

CA 2040180

An enzymatic process for the determination of .beta.-lactam antibiotics in a biological liquid, comprising the steps of (1) incubating the liquid with soluble D-alanyl-D-alanine-carboxypeptidase produced by Actinomadura R39, the .beta.-lactam antibiotic reacting with the enzyme to form an inactive and equimolecular enzyme-antibiotic complex; (2) incubating the mixture obtained with a solution of a thioester substrate having the formula Image wherein R1 = benzoyl, phenylacetyl or N .alpha.-acetyl-L-lysyl, R2 = glycyl or D-alanyl, and R3 = hydrogen or methyl so as to form, by hydrolysis, a 2-mercaptoalkanoic acid having the formula Image in an amount proportional to the residual enzymatic activity, said incubation being further conducted in the presence of glycine ar of a D-amino acid; (3) determining the amount of 2-mercaptoalkanoic acid formed, and (4) comparing the determination of step (3) with a standard, to obtain the antibiotic concentration in the biological liquid. A test set for carrying out this process is also disclosed.

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